Bulletin OEPP/EPPO Bulletin (2017) 47 (2), 164–173 ISSN 0250-8052. DOI: 10.1111/epp.12381 European and Mediterranean Plant Protection Organization Organisation Europe´enne et Me´diterrane´enne pour la Protection des Plantes PM 7/36 (2) Diagnostics Diagnostic PM 7/36 (2) Diabrotica virgifera virgifera Specific scope Specific approval and amendment This Standard describes a diagnostic protocol for Approved in 2003-09. Diabrotica virgifera virgifera. Revised in 2017-02. This Standard should be used in conjunction with PM 7/76 Use of EPPO diagnostic protocols. onto other flowering crops. Cucurbits are particularly attrac- 1. Introduction tive to corn rootworms, but these pests have also been The New World genus Diabrotica Chevrolat, 1836 is one found on lucerne, clover, rape, soybean and sunflower. of the largest leaf beetle genera, with about 354 described However, maize seems to be the preferred source of food species (Derunkov et al., 2015). Ten species or subspecies (Toepfer et al., 2015). within this genus are generally recognized as pests (Krysan Further information on the biology of Diabrotica can be & Miller, 1986); Diabrotica barberi Smith & Lawrence, found in the EPPO data sheet on Diabrotica barberi and Diabrotica undecimpunctata howardi Barber and Diabrotica virgifera (EPPO, 1997), information on geo- Diabrotica virgifera virgifera LeConte are serious pests of graphical distribution is available in the EPPO Global Data- maize in North America, the latter species in Europe too. base (EPPO, 2016). There are two subspecies of D. virgifera, virgifera (west- ern corn rootworm) and zeae Krysan & Smith (Mexican 2. Identity corn rootworm) (Krysan et al., 1980). Diabrotica virgifera virgifera is distributed from the Mid-Western to Eastern Name: Diabrotica virgifera virgifera LeConte and South-Eastern USA and northward into Ontario, Synonyms: Diabrotica virgifera Canada and is adapted to temperate climates (diapause), Taxonomic position: Insecta: Coleoptera: Chrysomelidae: while D. virgifera zeae is distributed mainly from Texas Galerucinae: Luperini: Diabroticina and Oklahoma in the United States to Panama and is EPPO Code: DIABVI adapted to warm climates (without diapause). Only the sub- Phytosanitary categorization: EPPO A2 List: no. 199, EU species D. virgifera virgifera has been found in the EPPO Annex designation: none.1 region. The pathways for introduction or spread of D. virgifera virgifera are infested soil containing eggs, lar- 3. Detection vae or pupae, or the aerial parts of maize plants (e.g. for fodder or green manure, or as cobs) carrying adults. In the Eggs of D. virgifera virgifera occur in the soil down to a latter case, however, the probability of spread with maize depth of 35 cm, most of them in the 15-cm layer (Baca plants is low because, before harvest, adults generally move et al., 1995) from late summer (August/September) and to attractive plants in other fields or soon die. Since it is throughout the winter as diapausing eggs. They occur in the likely that D. virgifera virgifera arrived by aeroplane, as soil of maize fields and, to a lesser extent, also in crops, shown by the fact that the first discovery site in Europe such as soybean and cereal, in neighbouring fields (Kiss was in close proximity to Belgrade airport (Edwards et al., et al., 2001). 1999), adults can also probably be transported as contami- nants on other means of transport (e.g. boats, trains, trucks, 1The EU decision 2003/766/EC on emergency measures to prevent the cars). Adult beetles can also fly up to 100 km from infested spread within the community of Diabrotica virgifera Le Conte was areas. Adult beetles are known to fly from mature maize repealed on 6 February 2014. 164 ª 2017 OEPP/EPPO, Bulletin OEPP/EPPO Bulletin 47, 164–173 PM 7/36 (2) Diabrotica virgifera virgifera 165 Fig. 1 Diabrotica virgifera virgifera, third larval instar. Larvae generally occur between May and early August Fig. 3 Pheromone trap. with a peak from May to June (Baca et al., 1995). There are three larval instars which live in the soil near traps should not be less than 20 m. They should also be maize roots. Grassy weeds (e.g. Setaria spp.) can serve as placed at points of entry, airports, ports and transhipment alternative larval hosts in maize fields (Moeser & Vidal, locations (including army barracks, if there is a risk of 2004). The highest number of larvae is found to a depth of movement of army equipment from infested areas) in order 15 cm (Baca et al., 1995). The main damage is caused to to monitor introductions. Host plant volatiles from both the root system. The third larval instar (Fig. 1) is mainly maize and cucurbit blossoms and some floral odorants responsible for most damage to the root system, and this attract adults (Hammack & Petroski, 2004). damage is commonly referred to as ‘root pruning’ (Fig. 2). European beetles that are commonly present in the vicin- Larvae pupate in soil chambers in the root zone. They ity of corn fields and that might be found in traps are pre- remain in the pupal stage for 7–10 days (Sivcev et al., sented in Fig. 4. Beetles that are regularly found in traps 2012). and that can be confused with D. virgifera virgifera when Adults of D. virgifera virgifera are found from the end in poor condition are presented in Fig. 5. of June to mid-October with a peak in the second half of July and during August. Adults most commonly occur on 4. Identification the leaf, tassel and silks (i.e. the leaf axil and ear tip of maize) before, during and after flowering, respectively Identification is usually based on morphology. Although (Baca et al., 1995). identification should be performed on adults, information is Pheromone traps (e.g. PALs) (Fig. 3) or yellow sticky also provided on eggs and larvae. traps are used to catch the adults. These should be placed in maize fields or nearby in June. Traps should be checked 4.1 Morphological identification at least every 2 weeks, but preferably each week, up to the end of September. The distance between the pheromone 4.1.1 Description of the genus Diabrotica The species in Diabrotica have an elongate, oval body, with the prothorax being narrower than the elytra (Derunkov et al., 2015). The basic colour of the beetles may be green or yellow, usually with spots, vittae or bands. The colour patterns in most Central American species are bright and variegated, and frequently iridescent. The head has short genae that are no longer than a quarter of the diameter of the eye. The antennae are long, filiform, rarely slightly ser- rate, and in males frequently modified, with the middle antennomeres being thickened or bearing a longitudinal costa on the inner side; antennomeres 2 and 3 are subequal in length, or antennomere 3 is slightly longer, but usually no more than 1.5 times longer than antennomere 2. The prono- tum is quadrate or subquadrate, usually with the ratio of width to length being 1.23–1.37; only rarely is the pronotum slightly transverse with the ratio of width to length exceed- Fig. 2 Diabrotica virgifera virgifera, ‘root pruning’ damage. ing 1.38. The pronotum is either with or without two discal ª 2017 OEPP/EPPO, Bulletin OEPP/EPPO Bulletin 47, 164–173 166 Diagnostics 3 mm Fig. 4 European beetles that are in the vicinity of corn fields (left to right): Cryptocephalus vittatus, Cryptocephalus decemmaculatus and Phyllobrotica quadrimaculata. Fig. 5 Beetles regularly found in traps that could be confused with Diabrotica virgifera virgifera when in poor condition (left to right): Demetrias atricapillus (Coleoptera: Carabidae), Xanthogaleruca luteola (Coleoptera: Chrysomelidae), Rhagonycha fulva (Coleoptera: Cantharidae). depressions; when present, the discal depressions may be (Lawrence, 1991). Observation should be made on the shallow, wide or deep, and they are sometimes represented third larval stage using a stereomicroscope with a mini- by a small, round fovea. The pronotum surface is smooth, mum magnification of 409 and several specimens should shining or shagreened. Shagreening may be represented be observed to take account of any polymorphism. A dis- by narrow mesh, minute wrinkles or minute tubercles. section of the cephalic capsule is necessary for examina- The elytra lack notches, although there are sometimes one tion of the mandibles. or two depressions on each elytron; they may be sulcate Chrysomelid larvae can be distinguished by the following with 2–5 distinct sinuate sulci, and they are usually dis- characters: tinctly shagreened. The elytral punctures are irregularly • mandibles palmate and toothed, without mola (Fig. 6) arranged. The legs are not modified. Males have apical • legs generally well developed, usually with 5 segments, spurs on the middle and posterior tibiae, but none on the including tarsus and pretarsus that are fused to form a anterior. Females have spurs on all tibiae. The front coxal tarsungulus (Fig. 7). cavities are open. Claws are bifid. The last ventral Larvae of subfamily Galerucinae can be identified by abdominal segment in males is slightly truncate, without combination of the following characters: an apical lobe. The aedeagus is symmetrical, or rarely • 2 antennal segments (Fig. 8) asymmetrical with a cavity on the right side, and it is • 0 or 1 pair of stemmata (Fig. 9) (no stemmata in without basal spurs; the orifice is covered by a sclerotized D. virgifera, Fig. 1) plate; and the internal sac has a complex armament con- • presence of 4–5 teeth on the mandibles (Fig. 6) (5 teeth sisting of robust, chitinized hooks or plates, with 2–6 for D. virgifera). sclerites. The larvae can be confused with other European soil- dwelling insect species (such as wireworms), which can be 4.1.2 Description of the species Diabrotica virgifera 4.1.2.1 Eggs.