483 13 11,12 b,d 8 ppm, . 0 Myzus per- ≥ (Razoumowsky), larvae in 2012 Society of Chemical Industry and alate morphs of c 6 ppm concentration. At and Daniel C Peck Gennadius (Hemiptera: Aleyrodidae). . 0 c ≥ B. tabaci Amphimallon majale Bemisia tabaci (Sulzer) (Hemiptera: Aphididae) preferred untreated leaves Department of Entomology, New YorkCornell State University, Geneva, Agricultural NY, Experiment USA Station, College of Life Sciences, Nankai University, Tianjin, China EntomoTech Fundamentals, Geneva, NY, USA Correspondence to: Gengping Zhu,Engineering, Nankai College University, Tianjin 300071, of China. E-mail: Environmental [email protected] Science and College of Environmental Science and Engineering,China Nankai University, Tianjin, Guoqing Liu Imidacloprid is a versatile insecticide that is used for systemic fo- ∗ c a d b b the outcome ofroot-feeding and soil-inhabiting insecticidal insect pest complexes interventionsglobal that agricultural are and against of economic concern. A better a understanding of how suite insecticides influence the of sublethal behaviorwill of lead soil to insects improved soil insect managementgreater via efficacy better and targeting, novel application approaches. liar and soil drench applications and as seedpelleting dressing, and film coating, multilayer coating,efficacious allowing protection for relatively of safe young and plants against insect attack. It has a relativelyeffect broad against spectrum early offects activity insect after and developmental exposure a stages. to long-lasting several imidacloprid Sublethal insect have ef- pests. been Disruption documentedobserved of for in host evaluation behaviorIn was choice tests, adult sicae Amphimallon majale L. ; movement arrestant; imidacloprid; sublethal effect; X-ray radiography Larvae Larvae (Balsamo) The mole 4,5 Giglio-Tos 3–10 6 Among white Popillia japonica Additionally, the 7,8 1 Agriotes obscurus Otiorhynchus sulcatus (Metchnikoff) Sorokin Amphimallon majale do not remotely detect imidacloprid in the soil; they neither evade contact, nor are repelled Scapteriscus borellii Beauveria bassiana Matthew J Petersen, This suggests that detection of ∗ Scudder modified their movement 15mininsoilmicrocosms. 9,10 . a,b A. majale ≤ L. Karst. (Pinales: Pinaceae). Among soil-inhabiting arthropod pests, : 483–492 www.soci.org (LeConte) were repelled by tefluthrin-treated 2 69 Metarhizium anisopliae 2013; M. anisopliae Picea abies white grub; European chafer;

Scapteriscus vicinus Limonius canus

2012 Society of Chemical Industry c and orientation to soil-borne antagonists are likely to influence Pest Manag Sci 1 INTRODUCTION The interrelated factors thatsoil influence insecticidal include activity chemistry in andsoil the formulation and of climatic the active conditions,target species ingredient, susceptibility and insect and developmental stage. behavior of the

Abstract BACKGROUND: Manipulative studies of theare behavioral stymied by response the difficulties of of soil-dwelling observing andout insects interpreting using interactions to played X-ray insecticides out radiography below and ground. to other Six quantify experiments antagonists the were carried movement of the European chafer, of the European chafer, Gengping Zhu, response to imidacloprid and how this was affected by host plantRESULTS: cues The and cold movement temperature. of third instars was arrested in imidacloprid-treated soil at (wileyonlinelibrary.com) DOI 10.1002/ps.3394 Imidacloprid as a contact arrestant for larvae Research Article Received: 29 September 2011 Revised: 30 June 2012 Accepted article published: 3 August 2012 Published online in Wiley Online Library: 28 August 2012 repellence behavior has been widely testedto and is insect highly relevant susceptibility and control product efficacy. presence of bothagents synthetic can insecticides and significantlymajor soil biological influence insect pest control complexes. the Understanding howand insecticides behavior neurotoxicants of modify insecthost-finding taxa behaviors, and from including feeding, mating, pest may offer management. new avenues for effective of the wirewormsand (Coleoptera: Elateridae) seeds after feedingof for the weevil (Coleoptera:(F.) were Curculionidae) significantlyhost deterred plant by bifenthrin absence of the crickets (Orthoptera: Gryllotalpidae) and (Hypocreales: Clavacipitaceae) and Vuillemin (Hypocreales:their Clavacipitaceae), exposure to ultimately the entomopathogenic reducing fungi. grubs (Coleoptera: Scarabaeidae), theNewman larvae of (Japanese beetle)particles avoid of soil that contains mycelial in response to the arrestant effect ofdisruptive imidacloprid effect was on stronger dispersal than distancedownward the in movement vertical attraction under a versus cue cold lateral posed temperature panels. treatment by In that germinating vertical simulated overwintering grass panels, conditions. thereCONCLUSION: seed. Larvae was There of a was less a disruptive less effect on after contact. Imidacloprid thereby actsthe synergistic as effect a of imidacloprid contact in arrestantoverwintering combination with behavior. to other disrupt biological agents grub for movement. white grub This control, finding and might its effects help on to grub explain Keywords: . et al 5cm . 0 : 483–492 × 5 69 . 2013; Pest Manag Sci The voltage was set at 74 kVp. C. ◦ 7,9,10,32 collected from three field sites in New York C until 24 h before the start of experiments when ◦ A. majale Diagrams of the PVC pipe and Plexiglas panel microcosms. they were transferred to 23 22.1 EXPERIMENTAL METHODS Source ofThird-instar insects State were usedwere to collected conduct on all 19Course, October laboratory Oswego 2009 County), experiments. on in 27 Grubs FultonFairways October Golf (Battle 2009 Course, in Ontario Island County) Victor Golf and (Parkview onRochester 9 November (Park 2009 Point in at RIT, Monroesoil, County). with Larvae a were piece held of in sodsource, from the at collection 4.4 site included as a food Figure 1. (A) PVC pipe with treated soilevasion before (106 contact g, with left) the treated and zone untreatedrepellence; would evasion soil be upon evidence (144 contact of g, would remote be right); evidence of(B) contact Plexiglas repellence. panel in a horizontalcircles (left) indicate and grub vertical (right) placement position; sites. the five 2.2 Radiographic imaging Non-destructiveradiographicimagesofinsectswithinmicrocosms were made with a Faxitron X-rayPackard, cabinet (Model Palo 43855B; Alto, Hewlett- CA). Thisinsect system behavior has been studies. widely used in soil mesh) sandy loam soilTwo (89.4% kinds sand, of 8.8% soil silt, microcosmpipes 1.8% were clay; (Fig. used: pH 1A) polyvinyl 7.5). chloride topanels (PVC) (Fig. measure 1B) to single measure grubwere multiple grub response taken response. and Radiographs ofpredetermined Plexiglas all time intervals. PVC Thethe pipes location microcosm of and each was Plexiglasas grub marked within panels distance on relative at the to regular, radiograph its original and release recorded point. A commercial A digital X-ray scannerGeraete system GmbH, Dickel, (Model Germany) EZ400; wasmanipulate the NTB used images. Eletronische to capture, read and 2.3 Experimental design Experiments were conductedprovide in an experimental soil chamberin microcosms for studying which designed grub the to movement soil moisture, could temperature be andall controlled. microcosms homogeneity Regardless contained of sterilized of the and treatment screened and (0 design, 18 19 With X-ray www.soci.org G Zhu larvae and to 36 20 Anomala Maladera 31 Compared Herein,use 2012 Society of Chemical Industry were chosen 32 Amphimallon 30 c A. majale 27 hasproventobe and greater over- increased suscepti- 21 Behavior modifications A. majale 22–25 The species occurs from 32 acousticmethods, Antifeedant effects were 29 35 A. majale larval behavior, a series of Reduced fecundity was docu- 14,15 While the alteration of insect 26 Monitoring soil insect behavior is andX-rayradiography. . Kuwayama (Hemiptera: Psyllidae), 15–17 . 37 33 (Marsham) (Coleoptera: Curculionidae) (Stål) (Hemiptera: Delphacidae). A. majale A. majale M. persicae and in winter wheat. 28 Diaphorina citri Non-destructive techniques useful for studying insects (Waterhouse)], control of (Arrow)], Japanese beetle and oriental beetle [ 34 (Razoumowsky) (Coleoptera: Scarabaeidae) has become Nilaparvata lugens To gain a better understanding of the nature of imidacloprid’s Since its first North American detection in 1940, for the followingpest status studies but notmaintained also because in only the the becausetarget developmental laboratory stage of of can for curative thethis be several control study species’ months in wasand and the orient to away is field. from determine imidacloprid,being the The (1) with considered: whether two evading first kinds larvae imidacloprid of objective beforetreated repellence could zone, contact of or with detect evasion the uponformer contact would with be the evidence insecticides. ofwould The remote be repellence, evidence while of the contactwere latter repellence. Subsequent to objectives determinelarval (2) what behavior concentrationsby response, cause the (3) changes how presencecauses in of the disruption host response toand plants grub is management and overwintering modified of (4)difficult whether behavior. root-feeding it imidacloprid The insects is study played is to out stymied access, below by ground. interpret how and manipulate interactions sublethal effects on experiments wasantagonists conducted and agonists, with namely soil-incorporatedand different imidacloprid germinating grass combinations seed. of Third-instar difficult and usually requires significanthabitat. alteration of the insect’s withinsoilincluderadioactivetagging, wileyonlinelibrary.com/journal/ps Ceutorhynchus obstrictus and over systemically treated leaves. behavior after exposure to imidaclopridfor has white been grubs, demonstrated repellenceexamined behavior under has controlled yetprid’s to widespread conditions. use be in Because thoroughly exploiting turf potential of environments, insect understanding imidaclo- behavioral and pellence modifications, behavior, such could as be re- insects, particularly important those for displaying low managementinsecticides. susceptibility of to contact soil regard to whiteof grubs, imidacloprid recent on studies oviposition have behavior, shown an impact orientalis difficult as it is less susceptible to insecticidal controls with the three otherin major the species of northeast turf-infesting Unitedcastanea white States, grubs Asiatic garden beetle [ wintering mortality of larvae. computedtomography(CT) environmental stressors such as drought. mented in bility to infection by entomopathogens a dominant whiteNew grub York pest State.nursery crops Elsewhere of it turfgrass has in caused many problems areas in of other majale due to insecticide exposureability may for control help of to explain the decreased is made of a non-destructive radiographic techniquewhether to imidacloprid determine influences the movement of through soil microcosms. the northeast Unitedpopulation States west now to established Michigan, in with British a disjunct Columbia. documented in

484 485 30 cm) C, 80% Schreb) ◦ × 25 × 5 . Two Plexiglas sheets were Festuca arundinacea 38 wileyonlinelibrary.com/journal/ps and Villani and Nyrop. 32 www.soci.org C, 80% RH) throughout the course of the experiment. Five ◦ For experiment 3, protocols were designed to measure how For experiment 4, protocols were designed to study movement For experiment 5, protocols were designed to study the effect of (0.2 g) was addedpositive in attractant. the The end seed ofpipe bottom was the before added adding pipe soil, by as orbefore settling by a adding inserting it food it to onto the source theafter the soil and rubber surface infestation stopper. (2, Pipes 6,destructively sampled were to 26, measure radiographed grub 50, mortality. It 74,that was predicted the 98, results 144 would andmovement vary would 266 be from h) arrested experiment after and 1seed. contact then with insofar the as germinating insect imidaclopridreducedthegrub’sresponsetograssseedunderano-choicesituation.Atotalof18replicatesoftwotreatments(36pipes)were used. Both treatmentswith had or grass without seed imidacloprid (0.5 (0.8was g) ppm). chosen The on on rate the one basis of side, being of just imidacloprid above the the threshold results of grub of response. The experimentswas opposite 1 side untreated and and 2 without as seed.infestation(2,6,26,50,74,98,144and266 Pipes were radiographed after sampled h)andthendestructively to measure grub mortality. It wasimidacloprid, predicted grubs that, would displace without towards the germinating seed, while with imidacloprid the displacement towards thebe seed would reduced. 2.3.2 Multiple-grub testsThe with design Plexiglas of panel the microcosms Plexiglas panel microcosmsand was based Wright on Villani of the 11Grass treatments seed (Kentucky-31 of tall experiment fescue, 1 (176 pipes) were used. separated and walled by a wooden frame (3 RH). After 24 h,grub one infestation according to pane each experiment. was Afterplaced the removed grub in was each to hole, the make Plexiglas was five reaffixed holes to the for panel. of multiple grubs in responsesoil(1700 to g)wasaddedtothebottomofthepanelandcompressed imidacloprid over time.to Treated a height of 15 cm.compressed to Untreated the soil top of (1700 the g) panel was for(Fig. an then 1B, additional added layer horizontal of and 15 position). cm throughout Grass the seed volume (4 of g) treatedthe was and grub mixed untreated with evenly soil a toThe food provide order source of over the fillingreduce course (treated of versus the the untreated) soil experiment. wasthree compaction treatments alternated [0 to (untreated bias. control), 0.4 Apanels) and were 2.0 total ppm] used. (12 The of total concentrationsa four were below- chosen and replicates to above-threshold represent of concentrationwere respectively. maintained Panels in a horizontal(28 position in the growth chamber larvae were introduced along thehorizontal position). midline All panels in were radiographed each at panel uniformintervals time after (Fig. infestation 1B, (24, 72 andgrub movement 144 would h). be It reduced at was 2.0 predicted ppm, but that not at 0.4 ppm. imidacloprid on vertical dispersal under overwinteringsimulated conditions by cold temperature. Untreated soil (2720to g) the was added bottom of the panelTreated and soil (680 compressed g) to was a then height added of and 24 compressed to cm. the top of (Fig. 1B) to hold athe uniform box amount was of removable soilraised to (3400 for 4% g). for both grub One treated and placement. pane untreatedin soil. of Soil The a panel vertical was moisture put position was fordetails filling of with experiments soil according 4,maintained to horizontally 5 the under and specific controlled 6. conditions Once (28 prepared, panels were ), C, 1 ◦ − 2012 Society of Chemical Industry c 443 g imidacloprid ha A. majale = ( 2 − : 483–492 69 2013; control is 17 mL 1000 ft Each pipe (diameter 2.5 cm, length 33.0 cm) (Fig. 1A) held 9 . For experiment 1, protocols were designed to ascertain the For experiment 2, protocols were designed to build on the Imidacloprid as a contact arrestant of a uniform amount ofhad soil a (250 hole g). (0.8 To cm introduceends. diameter) The grubs, on other each one edge edge, pipe of midway(0.4 the cm) between pipe evenly the had distributed ten along its smaller-diametermeasure length holes grub to serve location as relative markerssealed to to on the the release outside point.sealed with Holes with duct a were tape. rubber Eachpipe’s stopper end that of length filled the (2 an pipe cm).imidacloprid equal was solution Dry amount (for of treated soil soil) the orsoil) was tap to water raise added (for the untreated soil a moistureUsing to a certain plastic 12.5% funnel, before the amount pipe adding was to filled of throughtreated the two pipe. soil steps. Firstly, (106 g) was12 placed cm into with one a sideextended and wooden from compressed dowel. to the The rubbergrub treated stopper insertion to soil hole, 2.0 column or cmwas midline. thereby added distance and Secondly, from compressed untreated against the the soilto treated (144 accommodate soil, g) leaving the 2 rubber cm inserted stopper. into As untreated a soilcontrol and result, for 2 the cm compaction grub away bias, was from thepipe was treated order filled with soil. in soil treatments To which was alternated. eachpipes Once were prepared, side stored of horizontally under the controlled conditions (28 Pest Manag Sci formulation ofScience, imidacloprid Research Triangle Park, (Merit NC) was usedconcentrations to 2F; of prepare different soil Bayer microcosms. TheA. majale label Environmental rate of Merit 2F for 2.3.1 Single-grub testsThe with PVC pipe design microcosms ofet the al PVC pipe microcosms was based on Villani which approximates a concentration of 0.3loam ppm soil. for typical sandy threshold concentration ofgrub response, soil-incorporated and imidacloprid totime. examine for A changes in totalwere that used of response in over 12 thewith experiment. untreated replicates One-half soil, of while ofthe the 11 each 11 imidacloprid-incorporated other pipe soil treatments half was treatmentscontrol), was filled [0.0 (132 0.1, filled (untreated 0.2, with pipes) These one 0.4, concentrations represented of a 0.5, realistic range of 0.6, imidacloprid concentrations 0.8, in the 1.0, environment. 2.0, NoPipes grass 5.0 were seed radiographed was and included. after98 10.0 h). ppm]. infestation It was (2, predicted that, 6,grubs below would 26, some displace threshold 50, concentration, evenly 74 toand, the above and untreated some and threshold treated concentration,preferentially grubs sides, to the would untreated displace side. results of experimentobservations 1 over by a longer including time grass series. seed A total and of making 16 replicates 80% RH, photoperiod 12 : 12grub light : was dark). introduced After 24 at h,head the an in individual midline contact of with eachAfter untreated release, pipe each soil grub by was through placing observeddescend the and into its soil replaced within insertion if 10 it min. hole. To could prevent water not hole loss, the was insertion covered with masking tapeThe before pipes and were after then infestation. assembled3) in or sets eight of six (experiment (experimentsof 2), 1 the alternating and the treated left–right andand untreated orientation secured sides, with placed ducthorizontally on tape a for and Plexiglas radiography. at Plates plate, chamber random were throughout compass stored the course directionsremoved of in from experiment. controlled the conditions Pipes to were growth be only radiographed. . and 50% et al < 38 05). . : 483–492 0 while the 69 ≤ 40 P 1, C treatments. An 2013; ◦ = and Villani and Nyrop Pest Manag Sci 32 This approach is similar to the protocol 6 ppm, displacement was interrupted . 39 0 ≥ Owing to the small number of replicates, ANOVA 9 . 41 et al 6 position) of the treated side. Changes in the pattern of − 2 cm) and beyond was also compared using a chi-square test The disruption of displacement to the end of the treated In panel experiments, the distribution of grubs at each time ≤ assigned as the treated side. An ANOVA was alsowhether used to insecticide determine treatment affected the mean location ofrelative grubs to the release point in experimentwere 3. Orthogonal used contrasts to make aThis analytical separate approach comparison is for similar to eachin the time profile Villani analysis period. described side increased with imidaclopridconcentrations, concentration (Fig. 3). in At lower displaced particular along the entire length 0, of the pipe, whichwith 0.1, was an consistent uninterrupted search 0.4 for foodAt sources and (see concentrations experiment 0.5 2). ppm, of grubs 33.1 RESULTS Single-grub tests3.1.1 with PVC pipe microcosms Experiment 1 Grubs in thedifference untreated in controlintervals, displacement pipes confirming showed between thethe lack no sides experimental of significant a at protocolsproportion directional of (Fig. all 2). grubs bias displacing inherent Infor five to to all the the concentrations. untreated time There side treated wastreated was side significant pipes, for displacement all the to five time the time intervals intervals at 0.2 at and 5.0 10.0 ppm,for ppm, one for for time four two interval at time 1.0 ppm intervals (chi-square, at df 0.5 ppm and described in Villani and Wright was not used for experiments 1 and 2. interval was displayed visuallySigmaPlot 11.2 as software. a box plot constructed using ANOVA analyses were conducted usingSoftware. JMP Statistical Discovery effect of treatment on the distribution of grubs from( the center axis in experiment 4 (Fig.analyses 1B, horizontal were position). conducted All in the Microsoft chi-square Excel 2007, is useful forin revealing population the distribution rate overposition time. of relative to ANOVA movement the of release andimidacloprid the line on the grub mean was dispersal used grub change length to in4, the test panel the 5 in effect experiments and of 6.made between In the experiments untreated and 4groups. each In and two experiment 6, imidacloprid-treated the 5, comparisons were ANOVA madetwo between vertical comparisons the treatments and were between the two 28 after contact withend treated ( soil, and no grubs displaced to the 10 . www.soci.org G Zhu It was et al 38 2012 Society of Chemical Industry c C) (12 total panels) C treatments were ◦ ◦ larvae displacing to the untreated side away from a range of imidacloprid concentrations at 6, 26, 50, 74 and 98 h after C and horizontally at 28 A. majale ◦ C, 70% RH) during the course of the experiment. ◦ Percentage of C, vertically at 28 ◦ For experiment 6, protocols were designed to validate results of the previoustemperature experiment on and vertical furtherexperiment dispersal. 5, elaborate Panels but the all werezone role had prepared of an of imidacloprid-incorporated as untreated soil bottom in replicates (2.0 zone ppm). of A and three total a4.4 environmental of top four conditions (held vertically at All panels were radiographed at uniformplacement time (6, intervals 24 after and grub 48 h).movement It was would predicted that be downward(2.0 grub ppm) reduced concentrations. at low (0.4 ppm) but not high wileyonlinelibrary.com/journal/ps A significantly higher proportionuntreated of grubs side first was displacing considered(Fig. to 1A). evidence the Alternatively, for a significantly remotefirst higher repellence displacing proportion of to grubs theuntreated side, treated was side, considered andFor evidence for then the contact displacing untreated repellence. to control, the the side filled second (106 g) was introduction. An asterisk (*) indicates significant displacement to the treated side with respect to an even distribution (50%–50%). the panel for an additional layer ofGiven 6 the cm short (Fig. 1B, duration vertical of position). wasused.Atotaloffourreplicatesofthreetreatments[0(untreated the experiment (48 h), nocontrol), 0.4, grass and seed 2.0 ppm] (12were total introduced panels) along were a used. horizon Five 3(Fig. grubs cm 1B, from vertical the top position). ina each After panel vertical infestation, position panels in were aconditions kept growth (4.4 chamber in under cold temperature Figure 2. were used. All panels were radiographed atafter uniform time grub intervals placement (6,compared 24 in and 48 order h).the to Vertical disruption treatments test of were imidacloprid thepredicted on that effect downward downward dispersal length of movement. would be Itthe cold longer cold was under temperature temperature conditions. on The two 28 2.4 Statistical analysis Chi-square tests were used to test for differencesof in the proportion grubs distributedtube experiments, with the proportion respect ofwas grubs to compared in treatment the against untreatedbetween a and side sides null time. (50%–50%) expectation In approach of is for similar even to each distribution the treatment. profile analysis This described in analytical Fry predicted that dispersal from the release pointthe would vertical be position longer than in in the lateral position. compared in ordergrub to dispersal. Third-instar test grubscollected the used before effect in winter; the of onsetdevelopmental experiment stage pane of were elicits cold orientation a positive temperaturelarvae on geotropic during response tend where this to move downwards in the soil column.

486 487 62% -axis) y < wileyonlinelibrary.com/journal/ps 05). The proportion of grubs displaced to the . 6 to 6 axis represents the distance grub moved from 0 − ≤ P 1, = www.soci.org untreated; the The displacement of grubs was significantly inhibited by the = for all concentrations anddisplacement to time the treated intervals. side There forfor six was four time intervals significant time at intervals 2.0and ppm, at 10.0 ppm 0.5 and ppm, for forsquare, one df two time time interval at intervals 0.6 at and 1.0 5.0 ppm (chi- proportion of grubs displacing to the untreated side was untreated side tendedconcentration: to decline 25–60% with overconcentrations increasing (0–0.4 imidacloprid the ppm), seven 10–60%(0.5–1.0 time ppm) at and 5–40% intervals middle at at high concentrations concentrations (2.0–10.0 low ppm). presence of grass seed. In the untreated control pipes, grubs 2 − 4and − 2012 Society of Chemical Industry treated side, right side c = A. majale -axis: left side x -axis) ( z larvae to a range of imidacloprid concentrations and untreated control. Graphs indicate shifts in grub proportion ( : 483–492 A. majale 69 -axis) over time ( x 2013; Response of Imidacloprid as a contact arrestant of Pest Manag Sci grub distribution from 0.5, 0.6 to 0.8increasing ppm imidacloprid highlighted the concentration effect on of grubno distribution, apparent from effect to defined peaks at the positions of Figure 3. in microcosms ( the treated to the untreated side). 3.1.2 Experiment 2 Grub mortality was 25% in19 the to untreated 50% pipes, across and ranged differentthe concentrations from untreated in pipes the there treated were pipes.displacement no In between significant sides at differences all in seven time grub intervals (Fig. 4). The respectively (Fig. 3). Thedisplaced relatively to low the proportion untreated side ofarrestant might effect grubs be of that imidacloprid-treated due soil to (Fig. the 2). unexpected . 91, . et al 3 1, 95, 21 cm 42 cm. C, 45% . . ± ◦ 64 and 8 . 6 = : 483–492 4 6 . ± 02 cm at 6, ± . 69 1 ± . 7 2 . 1 ± . 49, df 073). Therefore, . . 6 2013; . 4 0 = = C the corresponding ◦ 08, 14 88 and 15 P . . F 19 and 4 . 5 7 C, by 48 h only 15% of 7 ◦ 96 and 6 ± ± . ± 6 5 1, 95, 4 . . 8 . ± Pest Manag Sci = 4 . 29, 14 . 19, 4 . 46 cm in the 2.0 ppm treatment. 7 . 6 64, 6 27, df . 5 . ± 6 3 ± 7 ± . ± 4 = . 5 1 . . F C treatments, the difference between the ◦ C was 6 ◦ 93 and 13 86 cm in the 0.4 ppm treatment and to 11 . . 4 6 05), but not at 0.4 ppm. Mean dispersal length in the . 0 ± ± 8 6 < For the two 28 . . values decreased to 4 horizontal and vertical treatment(Fig. of 8): grub 20 distribution and waszone 15% small respectively. However, of compared with grubs the grubs escaped distributed the imidacloprid-treated escaped to the untreatedlength at zone. 4.4 The mean downward dispersal grubs escaped the imidacloprid-treated zone, while, at 4.4 24 and 48 h respectively. In contrast, at 28 There was a marginalgrub significance mean effect position of relative to imidaclopridtreatments the (ANOVA, on release the line between the two At 0.4 ppm, grubsfollowed by were some arrested displacement intoand at the displaced untreated the in zone both release at directionsnot 72 point h, at displace 144 beyond at h. the 24 At releaseon h, 2.0 the points, ppm, axis and grubs at most did 72 were24 and h arrested 144 and h. displaced Only into three the untreated grubs zone. escaped the axis at 3.2.2 Experiment 5 In the untreatedfrom panels, the 85% release oftended point. grubs to In dispersed move therepresenting majority downwards grub treated downwards population was panels, shorter (Fig. inand 7B). the the 0.4 2.0 ppm The grubs ppm also box treatedeach time panels plot interval. than column Theto in shorter the column the and release untreatedlength. its point As position control concentration relative indicates increased at from the0.4 0 and (untreated decrease control) 2.0 in to ppm,a grub mean significant dispersal dispersal effect lengthto of shortened. the imidacloprid There release was on line mean at 2.0 position ppm relative (ANOVA, imidacloprid had a less disruptiveunder the effect cold on temperature treatment. downward dispersal P untreated panels was 13 at 6, 24 anddispersal 48 h respectively, length while, decreased in the to treated panel, 12 mean 14 12 Under low-temperature conditions, downward dispersaldisrupted was still by imidacloprid,dispersal but (experiment 4). to a lesser degree than lateral 3.2.3 Experiment 6 For the two vertical treatments, at 28 1, = www.soci.org G Zhu 05). The . 0 25, df . 2012 Society of Chemical Industry ≤ 11 0038). c . P 0 = 6 and 6 positions. 1, = F − P = 1, 255, 05). The proportion of grubs . = 0005; 98 h: . 0 0 ≤ = P larvae displacing to the untreated side away from a range of imidacloprid concentrations at 6, 26, 50, 98, 146, 194 and 6 and 6 positions when food was 52, df . P − 1, 8 = = F 1, 255, A. majale = 05), but not at earlier time intervals. Unlike the . 0 09, df 45% had displaced in the pipes at 0.1–0.5 ppm. At . 0009; 266 h: ≤ ≤ . 12 0 Percentage of P = = 2 and 6 positions after 98 h. After 266 h, no grubs had 1, F P − = 266 h after introduction. An asterisk (*) indicates significant displacement to the treated side compared with an even distribution (50%–50%). wileyonlinelibrary.com/journal/ps displaced to pipeend ends of within the experiment 50 h (Fig.without 5), food and in sources in contrast stayed experiment to 1 there the (Fig.94% 3). untreated until At of pipes 98 the h, grubs for instance, were at the Figure 4. present (Fig. 5),food but was only absent (Fig. 17% 3).displacement were Significant were found shifts with at in respect to thetime. both those At pattern concentration 0–0.5 and of positions ppm, grub two crests when appeared at the At 50 h, all grubs had displacedpipes, to the but end of the pipe in untreated low concentrations, displacement to the treatedand end was arrested delayed by thewere presence also of two imidaclopridthe in crests the at soil. 0.6–10.0 There ppm, but these appeared at 3.1.3 Experiment 3 Grub mortality was(Fig. 17 6A) and and treated (Fig. 25% 6B)pipes there pipes was after respectively. no significant In 266 effect the of h seedbetween untreated on in sides grub displacement at the 2, untreated and 6, 266 26, h 50, (chi-square, df 74 and 144 h, but there was at 98 displaced through the imidacloprid-treated soiland to none the pipe had end, Displacement turned to the back untreated side to wasthe reduced displace imidacloprid-treated after soil. contacting to the untreated side. displacing towards seed wasintervals. 66.7–83.3% In over the the treated lastin pipes five distribution there time between was sides adf at significant 98, difference 144 and 266 h (chi-square, untreated pipes where grubs displacedthe to treated seed, displacement pipes in was(Fig. 6). arrested Imidacloprid at treatments had thedistribution:significancewasobservedat50,98and266 a imidacloprid-treated significant soil effect on h(ANOVA,50 h: grub 255, 3.2 Multiple-grub tests3.2.1 with Plexiglas panel microcosms Experiment 4 In the untreated24 panels, h, grubs and continued displaced to displacesignificant in at difference 72 both in and displacement directions 144 between htime sides at (Fig. intervals. for There 7A), was all with a no three significant effectdistribution of treatments at on grub 72 and 144 h (chi-square, df mean location of grub position relative to thethe release line two between treatments and control was not significantly different.

488 489 treated side, = The observed 42,43 -axis: left side x -axis) ( z wileyonlinelibrary.com/journal/ps -axis)overtime( x larvae cannot remotely detect imidacloprid in the soil at www.soci.org 6 and 6 after 98 h. The 0.6–10.0 ppm concentration treatments show a − The behavioral response ofeither insect to pests to enhancetreatments antagonists by increasing or can or decreasing, to act respectively, theirwith contact diminish the treatment. the Thecan efficacy effect be of of chemicals classified insecticidal ovipositional), on attractant, repellent as insect or deterrent. behavior arrestant, stimulant (locomotor, feeding, 4 DISCUSSION grub morality was constant amongand treatments 3, in suggesting experiments that 2 theby movement the response toxicity was of notA. imidacloprid. majale caused The present results indicate that -axis) in microcosms ( y 19 and . 7 ± 08 cm at 6, 2012 Society of Chemical Industry . 8 . 2 c ± 5 19, 4 . . 6 ± 4 . A. majale 90 and 2 2and6after98h. . 1 − ± 4 . larvae to a range of imidacloprid concentrations and untreated control. Grass seeds added in both ends of the pipe serve 05). Therefore, imidacloprid had a less . 6 to 6 axis represents the distance grub moved from the treated to the untreated side). The control and the 0.1–0.5 ppm 0 − 86, 2 . < 1 : 483–492 P A. majale ± 69 4 . 1, 95, 2013; = untreated; the = Response of 42 cm.Themeanlocationofgrubsrelativetothereleaseline 66, df . . 6 8 ± = 2 . Imidacloprid as a contact arrestant of Pest Manag Sci just around the release pointgrubs in the escaped horizontal the treatment, release several vertical point treatment. and The mean moved dispersal downwardstreatment distance in in was the the 2 horizontal Figure 5. as food source for the grubs. Graphs indicate shifts in grub proportion ( 24 and 48 hthe respectively. corresponding values In increased to contrast, 4 in the vertical treatment right side concentration treatments show a similar pattern, with two crests at positions 4 similar pattern, with crests at positions between the two treatments was statisticallyF significant (ANOVA, disruptive effect on dispersal lengthin in the lateral the position. vertical position than . et al show 6 ppm . 0 : 483–492 ≥ Host plant 69 14 A. majale 2013; Pest Manag Sci did not detect imidacloprid in was also affected significantly by to available food sources could act The lack of significant differences in 13 B. tabaci B. tabaci A. majale with seed only, or seed and imidacloprid, right side grubs from treated areas and into surrounding = A. majale -axis: left side x The impact of imidacloprid on insect behavior has been little The attraction of prolonged contact with the chemicalwas indicates acting that as an imidacloprid arrestant. studied; however, it has been shown to elicit ain behavioral response select taxa.choice Female experiments using surface-treated leaf discs. imidacloprid when ingested, although it was notthrough similarly affected direct contact. evaluation behavior of untreated areas. to draw larvae into the presence of imidacloprid-treated soils, no preference for soilAlthough not it treated is or notfrom treated possible this with to work, imidacloprid. define it a is threshold apparent concentration that concentrations of directional movement indicates that larvae of arrest movement as earlyresults as 6 to h field after settings, contact.control the In of efficacy applying third-instar of these white imidacloprid grubbehavior, and for should any curative apparent be lack enhanced of control byrepellency. cannot be insect Furthermore, attributed to field applicationnot of expel imidacloprid would -axis) ( z C). ◦ www.soci.org G Zhu 2012 Society of Chemical Industry -axis) over time ( c x C and vertically at 4.4 ◦ larvae searching for food sources in untreated soil (A) and imidacloprid-treated (B) soil at 0.8 ppm. Graphs -axis) in microcosms ( A. majale y C, vertically at 28 ◦ 6 to 6 axis represents the distance grub moved from the left to the right side). − Box plot of grub population distribution in microcosms at 6, 24 Box plot of grub population distribution in Plexiglas panels in the horizontal (A) and the vertical (B) position. (A) Grub population in the treated Effect of imidacloprid on untreated; the = indicate shifts in grub proportion ( wileyonlinelibrary.com/journal/ps any of the tested concentrations,imidacloprid nor after do contacting larvae the move insecticide,at away even high from concentrations. when Furthermore, present lack of grub movement after Figure 8. and 48 h after grub placementconditions with (horizontally the at microcosms 28 held in three different (upper) and untreated (down)placement side in at the untreated 24, control 72 and and the surface 144 application h of after imidacloprid-treated soil grub at placement. 0.4 (B) and Grub 2.0 ppm population concentration. in vertical microcosms at 6, 24 and 48 h after grub Figure 7. Figure 6.

490 491 24 . and et al A. ma- :534–545 37 Metarhizium ¨ ofer :7–19 (2008). 89 :140–147 (2005). 34 Koppenh , and the dusky wireworm, :1441–1449 (2007). 36 Environ Entomol 22–25 25 . :149–168 (1988). Environ Entomol Heterorhabditis bacteriophera 33 Environ Entomol wileyonlinelibrary.com/journal/ps . Limonius canus , Soil application effects of A. majale :177–198 (1972). Environ Entomol et al :42–50 (2002). :1021–1029 (2006). (Coleoptera: Elateridae), to insecticide-treated 17 6 ppm. This finding might help to explain (Coleoptera: Elateridae), on exposure to synthetic . 48 larvae do not remotely detect imidacloprid 35 0 ≥ on Japanese beetle (Coleoptera: Scarabaeidae) behavior Annu Rev Entomol Beauveria bassiana Am Entomol www.soci.org Preston-Wilsey LM, anisopliae Agriotes obscures wheat seeds in a soil bioassay. responses of wirewormstreated wheat (Coleoptera: seed. Elateridae) to insecticide- Agriotes obscurus insecticides in a soil-less bioassay. (2008). the blackentomopathogenic fungus vine revealsEnviron Entomol weevil a new (Coleoptera: tritrophic interaction. Curculionidae) and an Brandenburg RL, Useobserving of mole radiography cricketsoil. (Orthoptera: and Gryllotalpidae) tunnel behavior castings in for crickets (Orthoptera: Gryllotalpidae)fungus, to the entomopathogenic Pacific Coast wireworm, Annu Rev Entomol behavior. movement is greatly reduced. This pattern may explain the 9 Villani MG, Krueger SR, Schroeder PC, Consolie F, Consolie NH, 4 van Herk WG and Vernon RS, Soil bioassay for5 studying van behavioral Herk WG, Vernon RS and Roitberg BD, Repellency of a wireworm, 6 Kepler RM and Bruck DJ, Examination of the interaction between 7 Villani MG, Allee LL, Preston-Wilsey LM, Consolie N, Xia Y and 8 Thompson SR and Brandenburg RL, Tunneling responses of mole 3 van Herk WG, Vernon RS, Moffat C and Harding C, Response of the 1 Harris CR, Factors influencing the2 effectiveness Haynes of KF, soil Sublethal insecticides. effects of neurotoxic insecticides on insect by sublethal rates of neonicotinoids. ACKNOWLEDGEMENTS The authors thank Dan Olmsteadgrub for collections, his valuable soil assistance preparations with Seto and statistical for analyses, technical Akiko support,protocols, Masanori Anuar Seto Morales for forand helpful developing comments Jim and early Opela, suggestions Richgrub Belowski collection and sites. David Major DamaskeScholarship support Council for was through access a provided to fellowship byand awarded by the to the Federal GZ China Formula and Funds GL, projects621819 NYG-621404 awarded and NYG- to DCP. ThisVillani paper (1953-2001) is and dedicated Haruo to Tashiro (1917-2009). the late Mike REFERENCES suggest that the major factortions responsible between for imidacloprid synergistic and interac- may entomopathogenic be nematodes thecauses general drastically disruption reduced of activitytheir of normal exposure the to nerve grubs, the function thusresult infective is increasing juvenile that in nematodes. line The withthat, present this when theory exposed and provides tojale empirical imidacloprid, evidence even atsynergistic interaction low between rates, imidacloprid for control of 5 CONCLUSIONS Amphimallon majale in the soil.contact. They Imidacloprid neither functions as evadegrub a contact, contact nor movement. arrestant to areconcentrations disrupt Movement repelled of after is arrested after contact with the synergistic effect ofbiological imidacloprid agents for in the control combination of late-instar with whitedisruption grubs, other of and grub the overwintering behavior by imidacloprid. A. 44 P. japonica 2012 Society of Chemical Industry c proposed that 26 grubs searching for . et al larvae after short-term A. majale The resulting processes may andthebehaviorpatternsof P. japonica 45 38 As such, the sluggish movement A. majale 38 has been reported previously, but direct : 483–492 69 2013; A. majale displayed here. The internal grub status interacting with the The capacity of imidacloprid to impact upon the overwintering Like other neonicotinoids, imidacloprid acts selectively on the Imidacloprid as a contact arrestant of emprical evidence is lacking. Grewal behavior of external factors determines their response tostimuli the present various in external the environment,microcosms. Within or soil as microcosms, shown larvae were herebe clearly within shown attracted to soil to available fooduntil sources, coming searching the into soil contact column some with grubs grass chose seed. the Duringresulting ‘right’ experiments, in direction their intogrubs successfully the chose reaching untreated the the side, ‘wrong’side, direction food resulting into source. in the a Other imidacloprid-treated search. reduction in By movement incorporating and imidacloprid,elicitors the arresting it movement cessation dominated of was those that revealed promotedsearch the for that food. Further understanding the the behavioralwhite response grubs of to agonists and antagonists might open opportunities to use their properties to retain grubs in, orzones attract in grubs to, some treated field conditions. majale insect central nervous system asnicotinic an acetylcholine agonist receptors. of the post-synaptic already stimulated by lowthe presence temperature of might imidacloprid, and be leadand to magnified potential a by exposure longer residence to time adversethese microcosm environmental experiments conditions. recorded response As after short-term exposure,notmorethan30 minbeforeholdingtheminthegrowthchamber, the extended exposure timesmightelicitastrongerresponseeffect.Additionally,whetherlarvae under natural conditions still reserve the capacity toexposure move to after imidacloprid, and a could more still be prolongedof released time by increasing the of temperature, stimuli is a subject of needed future study. include insect behavior modification.may Such lead changes to a in reduction behavior sure in to ecological biotic fitness and and abiotic increased mortalitygrub expo- factors. behavior, Modifications such of as white those shownsusceptible here, could to make biological them more controlsures agents. in this Coupling way control couldwhite mea- provide grub a management new approach insynergy for turf. between reduced-risk One control measures examplethird is of instars increased the to susceptibility entomopathogenic potential of nematodes when challenged Pest Manag Sci suggested that host plant location and acceptance by insectsresult is of the a shift in balanceand between external negative and inputs. positive The internal applicable shifting to balance published patterns analogy of is additionally thus facilitating greater utility of the product. Miller and Strickler germinatingseedasafoodsource imidacloprid reduces thelarvae survival of (third overwintering behavior. instar) The by result wasmortality altering attained, rate however, their of bythe third-instar normal fall comparing grubs and the overwintering 210 15controlled days days laboratory after after treatment conditions treatment indestructive and in the through X-ray next the spring. technology,displacement use Under were direct of made. non- observations Understudy, the the on application conditions of vertical of imidaclopridthe the did downward not present movement significantly deter exposure, of although within theit timeframe was of apparent these thatimpaired. experiments movement The third after instars used treatment in waspositive this geotropic experiment somewhat response have an and innate tend tocold move temperature downwards conditions. under . :3–9 et al 3 : 483–492 69 J Econ Entomol J Econ Entomol J Econ Entomol Ann Rev Entomol J Econ Entomol J Econ Entomol 2013; :1173–1182 (2000). . SAS Institute, Cary, :852–857 (2001). 93 57 Agric For Entomol :1–6 (1972). :1084–1098 (2008). (Coleoptera: Scarabaeidae) . (1960). 16 64 et al . SPSS, Inc., Chicago, IL (2009). :241–251 (1991). Turfgrass Insects of the United States Pest Manag Sci :102–119 (2006). 20 5 J Econ Entomol Pest Manag Sci :90–99 (2010). , ed. by Bell WJ and Carde RT. Chapman 66 . Microsoft Corporation, San Francisco, CA :132–144 (1988). Popillia japonica Pest Manag Sci 34 , Eavesdropping on insects hidden in soil and et al Environ Entomol . Cornell University Press, Ithaca, NY (1999). Pest Manag Sci New York’s Food and Life Sci Bull :2056–2060 (2009). :134–136 (1960). :1262–1266 (1996). :249–269 (1990). :438–442 (1955). :1714–1719 (1993). (2007). NC (2003). in terms of the53 responses they elicit from insects. chemicals in termsinsects: of the an locomotor102 update responses they of elicit from Dethier, Chemical Ecology of Insects and Hall, New York, NY, pp. 127–157 (1984). insecticide chemistry. diapause behavior and(Coleoptera: Scarabaeidae). survival of overwintering white grubs York. and other89 scarab larvae inand Canada nursery fields. insects in natural and managed systems. (2001). macroarthropod behavior in agricultural systems. 35 insecticides by tagging48 with radioactive cobalt. Lapointe SL, interior structures of plants. tomography studiesinstar of Pecan weevil86 the (Coleoptera: Curculionidae). burrowing behaviorJapanese of beetle fourth- microcosms. and European chafer grubs in soil–turfgrass Coleopterists Soc Monograph laboratory susceptibility of turf-infesting whiteScarabaeidae) grubs (Coleoptera: toproducts. biological, biorational and chemicalturfgrass infesting control scarab grub speciesBull (Coleoptera: Entomol Scarabaeidae). Soc Am JMP: The Statistical Discovery Software. Version 7 Microsoft Office Excel 2007 SigmaPlot for Windows. Version 11.2 41 42 Dethier VG, Barton BL and Smith CN, The designation43 of Miller chemicals JR, Siegert PY, Amimo FA and Walker ED,44 Designation Miller JR of and Strickler KL, Finding and45 accepting Jeschke P host and plants, Nauen in R, Neonicotinoids – from zero to hero in 28 Smitley DR, Incidence of 40 26 Grewal PS, Power KT and Shetlar DJ, Neonicotinoid insecticides alter 27 Tashiro H, The European chafer, a continuing lawn problem in New 29 Vittum P, Villani MG and Tashiro H, 33 Hunter MD, Out of sight, out of mind: the impacts of root-feeding 34 Villani MG and Wright RJ,35 Environmental Fredericksen CF and influences Lilly JH, Measuring on wireworm reactions to36 soil soil Mankin RW, Brandhorst-Hubbard J, Flanders KL, Zhang M, Crocker RL, 37 Harrison RD, Gardner WA, Tollner WE and Kinard DJ, X-ray computed 38 Villani MG and Nyrop JP, Age dependent39 movement patterns of 30 Jackson TA and Klein MG,31 Scarabs as Morales-Rodriguez pests: A, a continuing Ospina A problem. and Peck DC,32 Variation Villani MG in and Wright RJ, the Use of radiography in behavioral studies of Myzus www.soci.org G Zhu JEcon :169–180 :502–513 :287–293 :113–120 Diaphorina 51 23 88 Metarhizium Entomol Exp 105 . 2012 Society of Chemical Industry Ceutorhynchus Pest Manag Sci J Econ Entomol c :170–174 (2009). Pest Manag Sci 65 Biol Cont . :57–62 (1996). 48 L.) with the neonicotinoid Myzus persicae Environ Entomol with special reference to an Entomol Exp Appl . :145–153 (1995). :283–293 (2000). PestManagSci :101–108 (1999). . 44 Pestic Sci 94 24 . J New York Entomol Soc Popillia japonica Brassica napus Pflanzenschutz-Nachr Bayer (German edn) :870–877 (2009). Pestic Sci and Myzus persicae Bemisia tabaci 65 Myzus persicae and its modification by external or internal uptake of Physiol Entomol Nilaparvatalugens :431–439 (2007). Entomol Exp Appl :245–251 (2000). (Marsham) (Coleoptera: Curculionidae), to seed and (Deuteromycetes). 19 :249–255 (2002). 100 ¨ ¨ ¨ ofer AM, Grewal PS and Kaya HK, Synergism of imidacloprid ofer AM, Brown I, Gaugler R, Grewal PS, Kaya HK and ofer AM and Kaya HK, Synergism of imidacloprid and an 103 Pest Manag Sci . :618–623 (1998). :113–136 (1991). :1099–1105 (2008). :1329–1336 (2009). neonicotinoidinsecticidesforthecurativecontrolofthewhitegrubs Amphimallon majale (2009). and entomopathogenicmechanism. nematodes against white grubs: the Klein MG, Synergismimidacloprid against white grubs: of greenhouse andBiol field Cont evaluation. entomopathogenic nematodes and entomopathogenic nematode: a(Coleoptera: novel Scarabaeidae) approach control to white in grub turfgrass. 91 (1998). of imidacloprid on antifeeding response. sublethal effectsnicotianae of imidacloprid on nicotine-tolerant and survival in turfgrass microcosms. (1994). study of theScarabaeidae) to soil-incorporated mycelial response particles of ofanisopliae Japanese beetle larvae (Coleoptera: Entomol of imidacloprid onoviposition, Japanese egg beetle hatch, (Coleoptera: and Scarabaeidae) larval viability in turfgrass. potato leafroll virus transmissionAppl by 44 (1997). systemic insecticide. aspects of neonicotinoid uses64 in crop protection. Bemisia tabaci imidacloprid. of imidacloprid on sublethal effects of imidacloprid on Asian citrus psyllid, citri obstrictus treatments of canolacompounds ( clothianidin65 and imidacloprid. four insecticides on the reproduction and wingplanthopper, formation of brown 25 Morales-Rodriguez A and Peck DC, Synergies between biological and 24 Koppenh 23 Koppenh 15 Nauen R, Behaviour modifying effects of low systemic concentrations 16 Devine GJ, Harling ZK, Scarr AW and Devonshire AL, Lethal and 10 Fry RC, Fergusson-Kolmes LA, Kolmes SA and Villani MG, Radiographic 22 Koppenh 21 George J, Redmond CT, Royalty RN and Potter DA, Residual effects 17 Mowry TM and Ophus JD, Effects of sub-lethal imidacloprid levels on 12 Elbert A, Haas M, Springer B,13 Thielert W Isaacs R, and Cahill M and Nauen R, Byrne D, Host Applied plant evaluation behaviour of 11 Elbert A, Becker B, Harwtig J and Erdelen C, Imidacloprid – a new 14 Nauen R, Koob B and Elbert A, Antifeedant effects of sublethal dosages 18 Boina DR, Onagbola EO, Salyani M and Stelinski LL, Antifeedant and 20 Bao H, Liu S, Gu J, Wang X, Liang X and Liu Z, Sublethal effects of 19 Dosdall LM, Responses of the cabbage seedpod weevil, wileyonlinelibrary.com/journal/ps

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