ISSN: (Print) (Online) Journal homepage:

Journal of Natural History ISSN: 0022-2933 (Print) 1464-5262 (Online) Journal homepage: http://www.tandfonline.com/loi/tnah20 New species of Megastylus (Hymenoptera: Ichneumonidae: Orthocentrinae) reared from larvae of Keroplatidae fungus gnats (Diptera) in a Dutch orchid greenhouse Andrei E. Humala, H. Marjolein Kruidhof & Jozef B. Woelke To cite this article: Andrei E. Humala, H. Marjolein Kruidhof & Jozef B. Woelke (2017) New species of Megastylus (Hymenoptera: Ichneumonidae: Orthocentrinae) reared from larvae of Keroplatidae fungus gnats (Diptera) in a Dutch orchid greenhouse, Journal of Natural History, 51:1-2, 83-95, DOI: 10.1080/00222933.2016.1257074 To link to this article: http://dx.doi.org/10.1080/00222933.2016.1257074 2016 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group Published online: 14 Nov 2016. Submit your article to this journal Article views: 235 View related articles View Crossmark data Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalinformation?journalcode=tnah20 Download by: [Wageningen UR Library] Date: 13 June 2017, At: 07:50

JOURNAL OF NATURAL HISTORY, 2017 VOL. 51, NOS. 1 2, 83 95 http://dx.doi.org/10.1080/00222933.2016.1257074 New species of Megastylus (Hymenoptera: Ichneumonidae: Orthocentrinae) reared from larvae of Keroplatidae fungus gnats (Diptera) in a Dutch orchid greenhouse Andrei E. Humala a, H. Marjolein Kruidhof b and Jozef B. Woelke b a Forest Research Institute, Karelian Centre of Russian Academy of Science, Petrozavodsk, Russia; b Greenhouse Horticulture, Wageningen University & Research, Bleiswijk, The Netherlands ABSTRACT A new parasitoid wasp species belonging to the genus Megastylus (Hymenoptera: Ichneumonidae: Orthocentrinae) found in an orchid nursery in The Netherlands is described and illustrated: Megastylus woelkei sp. nov. It was reared from parasitized larvae of fungus gnats (Diptera: Keroplatidae). The biology of this new species and the possibility to use it in biological pest control are briefly discussed. http://zoobank.org/urn:lsid:zoobank.org:pub:7e405813-67c6-44cd-b068-97b79523592e ARTICLE HISTORY Received 20 March 2016 Accepted 31 October 2016 Online 14 November 2016 KEYWORDS Biological control; Lyprauta; parasitoid; Proceroplatus; orchid root damage Introduction Orchid nurseries in The Netherlands, especially those growing the hybrid cultivars of Phalaenopsis (Orchidaceae) and Cambria (Orchidaceae), are experiencing severe problems with root damage. Especially the root tips are affected, causing the roots to branch strongly just above the damaged area. This ultimately results in a reduced number of flower stems and a longer vegetative stage (Chandler and Pijnakker 2009; Pijnakker et al. 2010; Pijnakker and Leman 2013).Growers attribute this problem to the action of so-called potworms. These presumed worms are actually the larvae of fungus gnats (Diptera: Keroplatidae). They should therefore not be confused with the actual potworms, which are small earthworms from the family Enchytraeidae. In The Netherlands, three species of Keroplatidae have been found in orchid nurseries: Lyprauta chacoensis Edwards, Lyprauta cambria Chandler (Figure 3(d)) and Proceroplatus trinidadensis Lane (Chandler and Pijnakker 2009; Pijnakker et al. 2010). To our knowledge, these three species originate from the Neotropics, and within Europe they do not occur outside greenhouses (Evenhuis 2006; Chandler and Pijnakker 2009). The keroplatid larvae found in orchid nurseries in The Netherlands live between the roots and bark inside orchid pots. They produce acid webs (ph 2.7 or less) consisting of central tubes along which they move, as well as more net-like parts bearing numerous droplets of oxalic acid containing saliva that they use for catching and killing prey and for shelter (Matile 1997). During a survey of different keroplatid species as well as the general macrofauna found in orchid bark substrate among Dutch orchid nurseries, CONTACT Jozef B. Woelke joop.woelke@wur.nl Greenhouse Horticulture, Wageningen University & Research, P.O. Box 20, 2665 ZG Bleiswijk, The Netherlands 2016 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives License (http://creativecommons.org/licenses/by-nc-nd/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited, and is not altered, transformed, or built upon in any way.

84 A. E. HUMALA ET AL. several cocoons (Figure 3(b)) were found in a nursery compartment that was heavily infested with keroplatid larvae. These cocoons, which were similar in size but different in appearance to those of keroplatid pupae (Figure 3(c)), were allowed to emerge in cages. Several days later, instead of keroplatid adults, parasitoid wasps were observed inside these cages. Here we describe and illustrate the morphology of this newly found Megastylus Schiødte (Hymenoptera: Ichneumonidae: Orthocentrinae) parasitic wasp species. The biology of this new parasitoid and the possibility of using it as a biological control agent are briefly discussed. Orthocentrinae is a medium-sized subfamily of ichneumon wasps of approximately 500 described species with a worldwide distribution (Yu et al. 2012). Most of the known species are Palaearctic or Holarctic, whereas tropical faunas are largely understudied. Very little is known about the biology of Orthocentrinae. However, all reliable rearing records of Orthocentrinae refer to gnats from the superfamily Sciaroidea, and from some species it is known that they are koinobiont endoparasitoids of primitive Diptera larvae. These small-sized ichneumonids can be extremely numerous in fairly humid and shaded habitats, and samples from Malaise traps show that the Orthocentrinae can comprise up to 30% of all trapped ichneumon individuals (Gauld 1991; Humala 2003). Material and methods Since April 2015, several orchid nurseries in The Netherlands were visited for a survey on the presence of different keroplatid species as well as the general macrofauna of orchid bark substrate. On 29 June 2015 we received cocoons from an orchid nursery that were presumed to be keroplatid pupae. However, although similar in size, they were different in appearance from normal keroplatid pupae. On 2 July 2015 the first Megastylus wasps emerged from these cocoons. To confirm whether these wasps indeed developed from keroplatid larvae, new larvae were collected on 6 and 7 July 2015 at the concerned orchid nursery and were reared in the laboratory. These keroplatid larvae were put individually into plastic cups with a thin layer of agar on the bottom that kept several moistened pieces of bark in place, allowing the keroplatid larvae to hide and make a diffuse net. Carpoglyphus lactis L. (Acarina: Carpoglyphidae) mites were added as a food source, as previously collected keroplatid larvae had been observed to feed on mites and larvae of Sciaridae (Diptera). Cups were covered with a lid containing small holes and placed in a climate cabinet at 28 ± 1 C constant temperature, 80% relative humidity and a 16 : 8 h light : dark period. To prevent desiccation, a small amount of water was sprayed inside the cups whenever needed. Three times per week the keroplatid larvae were fed and cups were checked for the presence of Megastylus cocoons. The cocoons were subsequently placed on a piece of bark in an open Petri dish lined with moist cotton wool, and kept inside a cage at 21 C ± 1 C, 50 60% relative humidity and a 16 : 8 h light : dark period. Altogether, 25 specimens of Megastylus wasps emerged between 13 and 16 July 2015; all parasitoid wasps and empty cocoons were preserved in vials with 70% ethanol. Thereafter, the imagos of Megastylus were mounted, studied and described following the morphological terminology of Gauld (1991), with one minor change: the term temple was used to describe the upper part of the gena. Photographs of cocoons and adult wasps were taken at the Forest Research Institute KRC RAS with a DFC 290 digital camera attached to a Leica MZ9.5 stereomicroscope. Images were combined with Helicon Focus Pro (v.5.3) software.

JOURNAL OF NATURAL HISTORY 85 The specimens of the new Megastylus species have been deposited in the following collections: Natural History Museum, London, UK (BMNH); Zoological Institute of Russian Academy of Sciences, St Petersburg, Russia (ZISP); Forest Research Institute, Karelian Research Centre of Russian Academy of Sciences, Petrozavodsk, Russia (FRI); and Naturalis Biodiversity Center, Leiden, The Netherlands (RMNH). The holotype of the new Megastylus species is deposited at the Natural History Museum, London (BMNH). Results Altogether 13 females and 12 males of ichneumon wasps belonging to the genus Megastylus reared from keroplatid larvae were collected. Study of this material led us to conclude that this Megastylus species is new to science and its description is given below. Taxonomy Family ICHNEUMONIDAE Latreille, 1802 Subfamily ORTHOCENTRINAE Förster, 1869 Genus Megastylus Schiødte, 1838 Type species Megastylus cruentator Schiødte, 1838 Diagnosis The genus Megastylus can be easily separated from other congeners by the following characteristics: presence of large ovoid scape considerably inflated and obliquely truncate about 50 from transverse; long antennae and legs; absence of areolet in the fore wing of all described species; thin and short ovipositor hardly surpassing metasoma; first segment of metasoma fairly long and almost straight; basal flagellomere two or three times as long as following ones. As a rule, female flagellum is covered with long setae; tyloids on male flagellomeres are lacking. Megastylus woelkei Humala, sp. nov. (Figures 1 and 2) Diagnosis This species differs from other known Megastylus species by the combination of the following characters: small size; fore wing length 2.3 mm; female antenna with 32 35 flagellomeres, first flagellomere 4.5 times as long as wide; ocular ocellar distance 0.9 times maximum diameter of lateral ocellus; first tergite 2.3 times as long as apically wide, spiracles at anterior 0.4 of tergite, sternite at about 0.5 of tergite length; second tergite as long as apically wide, without longitudinal striae. From the related Holarctic Megastylus orbitator Schiødte, it can easily be distinguished by its smaller size, wider face, longer flagellum and malar space, comparatively short pubescence on the female flagellum, male face pale with a central dark spot, and the absence of pale markings along the frontal orbits in both sexes. Remarks: one undescribed Megastylus species from Mexico, of which only males are represented in the available collections, is similar to this species in face colour pattern

86 A. E. HUMALA ET AL. Figure 1. Megastylus woelkei sp. nov., holotype female: (a) whole insect, scale bar 0.5 mm; (b) head (anterior view), scale bar 0.2 mm; (c) head and mesosoma (latero-ventral view), scale bar 0.5 mm; (d) head (dorsal view), scale bar 0.2 mm; (e) propodeum and tergites 1 5 of metasoma (dorsal view), scale bar 0.5 mm; (f) wings, scale bar 0.5 mm. (with a central dark spot), but differs in yellowish inner eye orbits and pronotum, proportions and number of flagellomeres, weak occipital carina, more robust hind femur and larger size (fore wing 3.5 4.0 mm).

JOURNAL OF NATURAL HISTORY 87 Figure 2. Megastylus woelkei sp. nov., paratype male: (a) whole insect, scale bar 0.5 mm; (b d) variation of face colour (anterior view), scale bar 0.2 mm; (e) head (dorsal view), scale bar 0.2 mm; (f) parameres (lateral view), scale bar 0.2 mm. Description Female. Body length 3.4 mm, fore wing length 2.3 mm (Figure 1(a)). Head. Head as wide as high, nearly polished, sparsely punctate. Face at level of antennal sockets 0.7 times as wide as head width, its inner orbits slightly divergent

88 A. E. HUMALA ET AL. ventrally (Figure 1(b)). Antenna long, with 33 flagellomeres, first flagellomere obliquely truncate, about 4.5 times as long as wide, second flagellomere 1.8 times as long as wide; flagellomeres from eight quadrate; flagellum densely covered with long erect setae. Frons smooth, nearly polished; margins of antennal sockets somewhat raised; face scarcely punctate and covered with sparse long hairs. Clypeus strongly convex, about 2.0 times as wide as high, its apical margin truncate. Malar space long, 2.8 times basal width of mandible, 0.35 times as long as face width, with distinct subocular sulcus as coriaceous strip between eye and base of mandible (Figure 1(c)); anterior tentorial pits large and open. Mandible slender, strongly tapered and turned inwards, small lower tooth much shorter than upper tooth; maxillary palps long. Temples moderately long, 0.5 times transverse diameter of compound eye; occipital carina complete. Ocelli of moderate size, ocular ocellar distance 0.9 times maximum diameter of lateral ocellus, postocellar line 0.8 times as long as maximum diameter of lateral ocellus (Figure 1(d)). Mesosoma. Mesosoma coriaceous, 1.8 times as long as high; epomia lacking. Epicnemial carina well developed, distant from front edge of mesopleuron and ending at mid-height (Figure 1(c)). Mesoscutum convex, with sparse small punctures bearing long setae; notaulus shallow and indistinct. Scutellum with lateral longitudinal carinae reaching its posterior end. Sternaulus weak and indistinct. Propodeum with lateral longitudinal carina present, two transverse carinae weakly developed, and other carinae reduced (Figure 1(e)). Propodeal spiracle circular, separated from lateral longitudinal carina by one diameter of spiracle. Fore wing with cu-a slightly proximal to base of Rs&M by 0.2 times length of cu-a; vein 2m-cu with two bullae, areolet absent, hind wing with cu-a slightly reclivous, Cu1 depigmented, closer to 1A than to M (Figure 1(f)). Legs long; fore coxa with transverse basal carina (Figure 1(c)); hind coxa matt, hind femur 5.3 times as long as wide, 0.8 times as long as hind tibia; hind basitarsus 0.4 times as long as hind tibia; hind tibia somewhat banded in apical half with depression on its dorsal surface; tibial spurs straight and slender, tarsal claws slender and weakly curved. Metasoma. First tergite of metasoma slightly constricted behind spiracles, 2.3 times as long as apically wide, coriaceous; glymma lacking; spiracles at anterior 0.4 of tergite (Figure 1(e)); dorsal and lateral longitudinal carinae lacking; first sternite fused to tergite, about 0.5 times as long as tergite length, in lateral view forming weakly rounded lobe; tergite II coriaceous, as long as apically wide; thyridium small, close to anterior margin. Remaining tergites more finely sculptured towards apex; metasoma weakly compressed to apex; tergites covered with sparse long hairs. Sternites II V with sparse dark setiferous punctures, sclerites fuscous. Ovipositor straight and slender, its sheath as long as hind femur depth. Colour. Fuscous. Mandibles, except for reddish teeth, palps and tegula pale. Face brown, with light marks under antennal sockets. Scape and pedicel ventrally, first flagellomere, apical half of clypeus yellowish brown, rest of flagellum fuscous. Pronotum anteriorly and in hind corners, lower mesopleuron and mesepimeron reddish brown. Metasoma fuscous, tergite II brown with yellowish thyridium and apical margin; tergite III yellowish, more fuscous laterally; tergite IV with light brown medial spot

JOURNAL OF NATURAL HISTORY 89 anteriorly. Fore and mid legs yellowish; hind legs light brown. Hind tibia infuscate dorsally in apical 0.4. Wings hyaline, pterostigma and veins brown. Male. Similar to female. Body length 3.0 mm, fore wing 2.4 mm (Figure 2(a)). Head. Head width 1.1 times its height; head matt, sparsely punctate. Face at level of antennal sockets 0.55 times as wide as head width (Figure 2(b d)). Antenna long, exceeds body length, with 31 flagellomeres, first flagellomere about 5.7 times as long as wide, second flagellomere 2.8 times as long as wide; all flagellomeres longer than wide. Frons nearly polished, almost impunctate; face smooth, scarcely punctate. Clypeus convex, about 2.1 times as wide as high, apical margin nearly truncate. Malar space 2.5 times as long as basal width of mandible. Ocelli of moderate size, ocular ocellar distance as long as maximum diameter of lateral ocellus, postocellar distance 0.9 times as long as maximum diameter of lateral ocellus (Figure 2(e)). Mesosoma. Mesosoma length 1.7 times height. Legs long, slenderer than in females; hind coxa coriaceous, hind femur 6.0 times as long as wide, 0.7 times as long as hind tibia; hind basitarsus 0.45 times as long as hind tibia. Metasoma. First tergite of metasoma slender, somewhat constricted behind spiracles, 2.8 times as long as apically wide, coriaceous, with sparse long hairs laterally; spiracles protruding, situated at anterior 0.4 of tergite; post-petiole somewhat inflated in middle and narrowed to apex; first sternite 0.4 times as long as tergite length, in lateral view forming rounded lobe; second tergite 1.2 times as long as apically wide. Parameres strongly tapered to apex (Figure 2(f)). Colour. Colouration as in female except lighter on pronotum and hind femur; tergites III IV yellowish, more fuscous laterally; tergite V with light brown median spot along anterior margin. Head dark brown; face yellowish, except for fuscous lower corners and elongated central spot in upper part close to antennal sockets, more dusky on facial margins and malar space (Figure 2(b d)). Scape, pedicel and first flagellomere yellowish, rest of flagellum fuscous; mandibles except for reddish teeth, palps and tegulae pale. Mesosoma brown, mesepimeron and pronotum excluding anterior part and hind corners yellowish; propleuron mostly brown; metasoma fuscous, except for yellowish thyridium, parameres, and apical margins of tergite II, tergite III and tergites IV V anteriorly. Legs yellowish, fore and mid coxae and trochanters pale. Hind femur yellowish brown, hind tibia more fuscous with light basal band and dark brown area apically on dorsal face. Variation. Flagellum with 32 35 flagellomeres in females and 28 32 flagellomeres in males. Fore wing sometimes with single indistinct bulla in 2m-cu, hind wing with cu-a and abscissa of Cu1 between M and cu-a forming single arched vein; one female specimen with sternite I reaching level of spiracles only. Colouration of propleuron and mesopleuron varying from fuscous to reddish-brown in both sexes, usually mesosoma darker dorsally than ventrally. Some male paratypes pale yellow on entire tergite III. In some female paratypes clypeus entirely yellowish; face, marks along notauli and tip

90 A. E. HUMALA ET AL. of scutellum brownish. Hind femur and tibia range from yellowish to brown; male face varying from entirely pale to light brown; fuscous spot in centre of face sometimes enlarged as vertical mark connecting frons and clypeus (Figure 2(b d)), or sometimes totally reduced. Material examined Holotype female (BMNH), The Netherlands, Bleiswijk, 52.004771 N, 4.516682 E, orchid greenhouse, emerged 13 July 2015 ex larva of fungus gnat (Diptera: Keroplatidae) (leg. J. B.Woelke,H.M.Kruidhof and M.van Twist);Paratypes 12 females and 12 males (BMNH, ZISP, FRI, RMNH), same label, but dates of emergence 10 July 2015, 13 16 July 2015. Biology Koinobiont, solitary larval endoparasitoid. All specimens of this species were reared from keroplatid larvae (Figure 3(a)), which were found between the bark and roots in pots of the orchid hybrid cultivars of Cambria. Three species of Keroplatidae were recorded in the orchid nursery: Lyprauta chacoensis, Lyprauta cambria (Figure 3(d)) and Proceroplatus trinidadensis. Larvae of the new Megastylus species consume host tissues almost completely (Figure 3(a)). Cocoons are tightly woven; thin and semi-translucent, light brown in colour, 4.5 4.7 mm in length and 1.3 mm in diameter (Figure 3(b)). Distribution The Netherlands, orchid greenhouses. This species presumably originates from the Neotropics as its probable hosts also originate from there. Etymology The species is named after its first collector and co-author of this article Jozef B. Woelke. Discussion Orchid growers face problems with root damage, especially damage to the root tips that results in increased branching of the roots just above the affected area. They attribute this damage to the action of Keroplatidae larvae, based on the frequent presence of a keroplatid larva in the vicinity of damaged roots (Chandler and Pijnakker 2009; Pijnakker et al. 2010; Pijnakker and Leman 2013). The biology of keroplatid larvae is still largely unknown. All known species can be divided into two groups: larvae that are predaceous, spinning toxic diffuse nets with a low ph in order to catch prey, and larvae that are fungal spore feeders, spinning a sheet-like web to catch spores of polyporous fungi (Aiello and Jolivet 1996; Matile 1996, 1997). The keroplatid larvae found during this survey all spun diffuse nets. It was also observed that they were actively hunting in the rearing cups when Carpoglyphus lactis mites and Sciaridae larvae were added. Even though the occurrence of orchid root damage seems to be related to the presence of keroplatid larvae, these larvae have not been observed to be actively feeding on the roots. The diet of keroplatid larvae as well as the exact relationship between the keroplatid larvae found in orchid nurseries and orchid root damage require further investigation.

JOURNAL OF NATURAL HISTORY 91 Figure 3. (a) Keroplatid larva parasitized by Megastylus woelkei sp. nov., inside the keroplatid larva the larva of M. woelkei is visible; (b) cocoon of M. woelkei sp. nov. from which the adult wasp has already emerged, scale bar 1.0 mm; (c) keroplatid pupa from orchid nursery, scale bar 1.0 mm; (d) female of Lyprauta cambria, one of the three probable host species for M. woelkei sp. nov., scale bar 1.0 mm.

92 A. E. HUMALA ET AL. The three keroplatid species found in Dutch orchid nurseries: Lyprauta chacoensis, Lyprauta cambria (Figure 3(d)) and Proceroplatus trinidadensis, are not of Dutch origin (Evenhuis 2006; Chandler and Pijnakker 2009; Pijnakker et al. 2010). From each of the genera Lyprauta Edwards (Diptera: Keroplatidae) and Proceroplatus Edwards (Diptera: Keroplatidae) only a single species occurs in the Palaearctic region (Evenhuis 2006). Twenty-six species of the genus Lyprauta have been described worldwide, of which nine species are found in the Neotropics including L. chacoensis (Evenhuis 2006). Of the genus Proceroplatus, 37 species are known worldwide, and 24 species are found in the Neotropics including P. trinidadensis (Evenhuis 2006). The exact origin of L. cambria is unknown, because this species was first discovered in an orchid nursery in The Netherlands. However, this species is also believed to originate from the Neotropics (Chandler and Pijnakker 2009). As previously mentioned, three keroplatid species occur in the orchid nursery where Megastylus woelkei sp. nov. was found. Because the larvae of all these species look very similar (Figure 3(a)), they could not be separated and identified to species level, and all of them could be considered as a suitable host. The most abundant keroplatid species recovered from a light trap in the orchid nursery where M. woelkei was first discovered was L. chacoensis, followed by L. cambria. Also, a single specimen of P. trinidadensis was recovered. It seems likely that all keroplatid species were parasitized, because at the time wasps were abundant in the greenhouse almost no adult keroplatids were monitored in the traps. As a consequence, the wasp population also declined. The cosmopolitan genus Megastylus Schiødte (Hymenoptera: Ichneumonidae: Orthocentrinae) comprises 35 described species and is divided into two subgenera: Megastylus with 27 species and Dicolus Förster with eight species (Yu et al. 2012; Humala 2014). However, only the Holarctic fauna has been studied, and many species of this genus are not described yet. Little is known about the hosts of Megastylus species but all known hosts of Megastylus species are restricted to the family Keroplatidae. Published host records are: Megastylus caseyi Ashmead and Megastylus orbitator Schiødte in North Carolina, USA, were reared from Orfelia inops Coquillett (Townes 1945; Dasch 1992); Megastylus amoenus Dasch was reared from Macrocera sp. in Tennessee, USA (Dasch 1992); Megastylus cruentator Schiødte was reared from Platyura sp. (Mansbridge 1933); Megastylus panamensis Wahl was reared in Panama from Orfelia billuus Matile (Wahl 1996). The distribution of M. woelkei in Dutch greenhouses is not yet fully known. From the 10 orchid nurseries that were included in the survey, M. woelkei was only detected in a single nursery. However, approximately 8 months after the first discovery of M. woelkei this parasitoid was also detected in another Dutch orchid nursery that had not participated in the survey. Interestingly, this second nursery has not recently exchanged any plant material with the first nursery. Moreover, the origin of M. woelkei is unknown and difficult to verify. The most likely option is that it has the same origin as the Keroplatidae host species, which are all believed to originate from the Neotropics (Evenhuis 2006; Chandler and Pijnakker 2009). How exactly these keroplatid species arrived in The Netherlands is unclear. They may have arrived with orchid plant material imported from the Neotropics. However, they could have been introduced with other ornamental species as well, as apart from Dutch orchid greenhouses keroplatids have also been found to occur on Anthurium (Araceae), Gerbera

JOURNAL OF NATURAL HISTORY 93 (Asteraceae) and other potted plants in Dutch greenhouses (Pijnakker et al. 2010). It may therefore well be that plant material imported to The Netherlands from the Neotropics also contained keroplatid larvae that were parasitized by M. woelkei. Moreover, the chance of discovering a new Megastylus species that originates from the Neotropics is relatively high. The West Palaearctic Megastylus species are rather well known (Förster 1871; van Rossem 1974, 1983), but the Neotropical fauna of the genus Megastylus, as well as all orthocentrines, are still mostly unstudied (Veijalainen et al. 2012, 2013). Until now, only two Neotropical species of Megastylus have been described: Megastylus panamensis from Panama (Wahl 1996) and Megastylus kasparyani Humala from Mexico (Humala 2014), and the number of Megastylus morphospecies registered in the Neotropics is higher than the number of described species in the world (Veijalainen et al. 2012). Hopefully, future studies of the Neotropical fauna will allow the newly described species to be found in its natural habitat and its origin can be clarified. Other possible sources of origin of M. woelkei seem less plausible, for the reasons outlined hereafter. One possibility would be that parasitized keroplatid larvae were brought to the greenhouse together with substrate imported for use in orchid pots. The three most frequently used substrates (in mixtures) are bark, coco chips and sphagnum. Bark used in Dutch orchid nurseries predominantly originates from Portugal and Spain, and bark from these two countries is commonly treated against insects by steaming. Therefore, the chance that M. woelkei has arrived with Portuguese or Spanish bark is relatively low. A small proportion of the bark is from Sweden. Although Swedish bark is untreated and therefore could contain parasitized keroplatid larvae, the chance of discovering a new Megastylus species from Sweden is rather low because of the relatively well-studied Swedish Megastylus fauna (van Rossem 1983). Coco chips are imported from Sri Lanka and are dried and pressed into blocks before transport, making survival of insects in this substrate unlikely. Sphagnum, which is imported from Chile, is also dried and pressed before transport, and is irradiated upon arrival to kill plant seeds. Another theoretical possibility would be that M. woelkei occurs locally in The Netherlands, entered the orchid nurseries through windows that are opened on warm days, and effectively located keroplatid larvae to parasitize. However, this scenario seems rather unlikely. Moreover, the Dutch Megastylus fauna is well studied (van Rossem 1974, 1983), so the chance of discovering an unknown species here is relatively low. So far, no effective biological control agent against keroplatid larvae has been identified. Adults of Dalotia (formerly Atheta) coriaria Kraatz (Coleoptera: Staphylinidae), the predatory mites Macrocheles robustulus Berlese (Acari: Macrochelidae), Hypoaspis miles Berlese and Hypoaspis aculeifer Canestrini (Acari: Laelapidae) and the entomopathogenic nematodes Steinernema feltiae Filipjev (Nematoda: Steinernematidae) and Heterorhabditis bacteriophora Poinar (Nematoda: Heterorhabditidae) are not sufficiently effective. They do not persist in bark substrate and/or do not attack the larger keroplatid larvae, which may be due to the oxalic acid secreted by these larvae (Pijnakker and Ramakers 2010; Pijnakker and Leman 2013). Megastylus woelkei may be used as a biological control agent against keroplatids in orchid nurseries as their presence was preceded by a strong decline in keroplatid adults in the greenhouse where M. woelkei was first discovered. More research is needed on the biology of Keroplatidae, on rearing methods of Megastylus wasps, and on the effectiveness of M. woelkei as a biological control agent against keroplatids in orchid nurseries.

94 A. E. HUMALA ET AL. Acknowledgements We would like to thank Marcel van Twist for his help with collecting material for this study. Sarah van Broekhoven is thanked for comments on an earlier version of the manuscript. Disclosure statement No potential conflict of interest was reported by the authors. Funding This project has been funded by Anthura BV as well as by a Public Private Partnership (PPS) Project (3742202500), that was in turn funded by the Dutch Ministry of Economic Affairs, the Dutch Product Board for Horticulture, the Potted Orchids Cooperative (an initiative by Dutch growers) and Horticoop BV. References Aiello A, Jolivet P. 1996. Myrmecophily in Keroplatidae (Diptera: Sciaroidea). J New York Entomol S. 104:226 230. Chandler PJ, Pijnakker J. 2009. Tropical fungus gnats established in nurseries in The Netherlands (Diptera: Keroplatidae and Mycetophilidae). Br J Ent Nat Hist. 22:81 93. Dasch CE. 1992. The Ichneumon-flies of America north of Mexico: part 12. Subfamilies Microleptinae, Helictinae, Cylloceriinae and Oxytorinae (Hymenoptera: Ichneumonidae). Mem Am Entomol Inst. 52:1 470. Evenhuis NL. 2006. Catalog of the Keroplatidae of the world (Insecta: Diptera). Bishop Mus Bull Entomol. 13:1 178. Förster A. 1869. Synopsis der Familien und Gattungen der Ichneumonen. Verhandlungen des Naturhistorischen Vereins der Preussischen Rheinlande und Westfalens. 25:135 221. (In German). Förster A. 1871. Übersicht der Gattungen und Arten der Familie der Plectiscoiden. Verhandlungen des Naturhistorischen Vereins der Preussischen Rheinlande und Westfalens. 28:71 123. (In German). Gauld ID. 1991. The Ichneumonidae of Costa Rica, 1. Mem Am Entomol Inst. 47:1 589. Humala AE. 2003. The ichneumonid wasps in the fauna of Russia and adjacent countries: Microleptinae and Oxytorinae (Hymenoptera: Ichneumonidae). Moscow: Nauka; p. 175. (In Russian). Humala AE. 2014. Mexican species of the subgenus Dicolus Förster, genus Megastylus Schiødte (Hymenoptera: Ichneumonidae: Orthocentrinae), with descriptions of a new species. Proc Russian Entomol Soc. 85:98 102. Latreille PA. 1802. Histoire naturelle, générale et particulière, des Crustacés et des Insectes. Tome 3. Paris: F. Dufart; p. 468. (In French). Mansbridge GH. 1933. On the biology of some Ceroplatinae and Macrocerinae (Diptera, Mycetophilidae). T Roy Ent Soc London. 81:75 92. Matile L. 1996. A new neotropical fungus gnat (Diptera: Sciaroidea: Keroplatidae) with myrmecophagous larvae. J New York Entomol S. 104:216 220. Matile L. 1997. Phylogeny and evolution of the larval diet in the Sciaroidea (Diptera, Bibionomorpha) since the Mesozoic. In Grandcolas P. editor. The origin of biodiversity in insects: phylogenetic test of evolutionary scenarios. Mém Mus Natn Hist Nat. 173:273 303. Pijnakker J, Leman A. 2013. Biological and chemical control of Lyprauta spp. in Phalaenopsis. Report GTB-1236. Wageningen UR Horticulture, Bleiswijk; p. 1 22. (in Dutch). Pijnakker J, Ramakers P. 2010. Lyprauta spp. (Diptera: Keroplatidae) in orchid greenhouses in The Netherlands. Miami (FL): Wageningen UR Horticulture.

JOURNAL OF NATURAL HISTORY 95 Pijnakker J, Ramakers P, Leman A, Ludeking D. 2010. Inventory of diptera larvae in greenhouses. Report GTB-1018. Wageningen UR Horticulture, Bleiswijk; p. 1 22. (in Dutch). Schiødte G. 1838. Ichneumonidarum ad faunam Daniae pertinentium, genera et species novae. Revue Zoologique par la Société Cuvierienne. 1:139 141. (In Latin). Townes HK. 1945. A catalogue and reclassification of the Nearctic Ichneumonidae (Hymenoptera). Part II. The subfamilies Mesoleiinae, Plectiscinae, Orthocentrinae, Diplazontinae, Metopiinae, Ophioninae, Mesochorinae. Mem Am Entomol Soc. 11:478 925. van Rossem G. 1974. The Gravenhorst, Schiødte and Foerster types belonging to the genus Megastylus Schiødte, 1838, with keys to the species (Hymenoptera, Ichneumonidae, Microleptinae). Tijdschr Entomol. 117:273 285. van Rossem G. 1983. A revision of Western Palaearctic Oxytorine genera. Part 4. Genus Megastylus (Hymenoptera, Ichneumonidae). Entomofauna. 4:121 132. Veijalainen A, Sääksjärvi IE, Erwin TL, Gómez IC, Longino JT. 2013. Subfamily composition of Ichneumonidae (Hymenoptera) from western Amazonia: insights into diversity of tropical parasitoid wasps. Insect Conserv Diver. 6:28 37. Veijalainen A, Wahlberg N, Broad GR, Erwin TL, Longino JT, Sääksjärvi IE. 2012. Unprecedented ichneumonid parasitoid wasp diversity in tropical forests. P Roy Soc B. 279:4694 4698. Wahl DB. 1996. Two new species of Megastylus from the new world (Hymenoptera: Ichneumonidae; Orthocentrinae). J New York Entomol S. 104:221 225. Yu DSK, Van Achterberg C, Horstmann K. 2012. Taxapad 2012, Ichneumonoidea 2011. Database on flash-drive. www.taxapad.com, Ottawa, Ontario, Canada [cited 2015 Dec 8]. Available from: www.taxapad.com