EFFECTS OF GRASSHOPPER-CONTROL INSECTICIDES ON SURVIVAL AND BRAIN ACETYLCHOLINESTERASE OF PHEASANT (PHASIANUS COLCHICUS) CHICKS

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1 Evirometl Toxicology d Chemistry, Vol. 5, No., pp. 58 5, SETAC Prited i the USA 7-78/9 $.. EFFECTS OF GRASSHOPPER-CONTROL INSECTICIDES ON SURVIVAL AND BRAIN ACETYLCHOLINESTERASE OF PHEASANT (PHASIANUS COLCHICUS) CHICKS PAMELA A. MARTIN,* DANIEL L. JOHNSON d DOUGLAS J. FORSYTH RR 8, Site, Box 9, Lethridge, Alert TJ P, Cd Reserch Cetre, Agriculture d Agri-food Cd, Lethridge, Alert TJ B, Cd Cdi Wildlife Service, 5 Perimeter Rod, Ssktoo, Ssktchew S7N X, Cd (Received Mrch 995; Accepted August 995) AstrctRig-ecked phest (Phsius colchicus) chicks were exposed vi cotct d/or igestio to formultios of three isecticides (Lors E, chlorpyrifos; Cygo 8E, dimethote; d Furd 8F, crofur) pplied to psture plots t oe d four times the rte recommeded for cotrol of grsshoppers (Orthopter: Acridide) i pririe Cd. Chicks ( d old) were exposed for 8 h i pes with the spryed vegettio d were fed either uspryed grsshoppers or grsshoppers spryed t the sme rtes s the vegettio. Cotrol groups were exposed to uspryed vegettio d received uspryed grsshoppers. Three replictes were coducted throughout Jue d erly July 99. Although some sigs of cetylcholiesterse (AChE) depressio were oserved i chicks exposed to isecticides, there ws o differece i the umer of mortlities mog tretmet d cotrol groups. Chicks tht died durig the exposure period gied sigifictly less weight th survivors i ll groups. Amog survivig chicks, weight gis of those exposed to the high- rte Furd tretmet cosumig spryed food were sigifictly lower th those of cotrols (.8 vs.. g/d). Bri AChE ctivity ws lower overll i survivig chicks th i those tht died; it ws ot sigifictly reduced mog chicks tht died i y tretmet group. Overll, survivors of Furd- exposed tretmet regimes hd lower AChE ctivity th those of Cygo d Lors tretmets; irds i high-spry-rte tretmets of ll isecticides hd lower AChE ctivity th those i ll low-spry-rte tretmets; d irds i ll tretmets cosumig spryed grsshoppers hd lower AChE ctivity th those fed uspryed grsshoppers. Food cosumptio ws ot ffected y y tretmet. KeywordsAcetylcholiesterse Crofur Chlorpyrifos Dimethote Riged-ecked phests INTRODUCTION I yers of severe grsshopper ifesttios, lrge res of the grssld d cerel cropld i wester Cd re spryed with isecticides. I 985, etwee.7 d.7 millio h cross the three pririe provices were spryed t lest oce; i 98, etwee.8 d. millio h were spryed i Ssktchew loe [,]. Up to % of the rle ld re i Alert hs ee ifested sice 98, d frmers spet $ millio o grsshopper-cotrol isecticides durig 985 d 98 []. Becuse grsshopper eggs overwiter i ucultivted res such s pstures, rgeld, hyfields, d rodsides, these res re preferetilly spryed s the ymphs emerge i the sprig. Such res frequetly represet the oly remts of wildlife hitt ville i the itesively frmed pririe ldscpe. Mgers of coopertive d privte grzig ld ttempt to cotrol grsshoppers o their property uder pressure from djcet crop frmers d i effort to limit the reductio i grzig potetil d forge productio o psture vegettio or hyfields. Thus, wildlife is t risk of exposure to grsshopper-cotrol isecticides over widespred res durig yers of grsshopper ifesttio d i more loclized loctios etwee outreks. Liquid formultios of the choliesterse-ihiitig orgophosphorus d crmte compouds re the isecticides most commoly used i recet grsshopper-cotrol progrms. Crofur typiclly ccouts for the mjority of the isecticide mrket. This isecticide hs ee reported to cuse deth i wterfowl feedig i spryed vegettio o witerig grouds [] d i gulls (Lrus cliforicus) gorgig o grsshoppers i spryed fields i Ssktchew [5]. I 985 d 98, di- * To whom correspodece my e ddressed. methote ws implicted i the deths of lrge umers of sge grouse (Cetrocercus urophsius) i lflf fields i Idho []. Such reports suggest tht these two isecticides my preset more widespred hzrd to grssld irds. Previous studies hve idicted tht mllrd (As pltyrhychos) duckligs wlkig to m through upld vegettio spryed with either crofur, dimethote, or chlorpyrifos were essetilly uffected uless feedig occurred e route [7,8, P.A. Mrti d D.J. Forsyth, upulished dt]. Juveile wterfowl sped very little time feedig i the uplds, wheres glliform species, which re permet residets of upld hitt, re proly t greter risk of pesticide effects. Shrp-tiled grouse (Tympuchus phsiellus), rigecked phests (Phsius colchicus), d gry prtridge (Perdix perdix) use pstures, hyfields, field mrgis, d rodsides extesively durig the rood-rerig period [9 ]. Spryig for grsshoppers i these res occurs from lte My through lte July, coicidig with the rood-rerig period. Very youg chicks my e prticulrly susceptile to direct toxic effects of ticholiesterse isecticides ecuse of their smll ody size. They my lso e more sesitive to isecticides so tht smller dose reltive to ody weight could cuse itoxictio, lthough the opposite hs lso ee demostrted [,]. Durig their first to weeks of life, juveile shrp-tiled grouse, gry prtridge, d phests cosume lrge qutities of rthropods [9,,], food source tht my e cotmited y isecticides followig spry pplictios. Choliesterse-ihiitig isecticides iterfere with thermoregultio i irds d my decrese the ility of youg chicks, which hve icompletely developed thermoregultory systems, to miti ody temperture durig periods of cold, wet wether [5,]. 58

2 Effects of isecticides o phests Eviro. Toxicol. Chem. 5, The ojectives of this study were to ssess the impct of three isecticides commoly used for grsshopper cotrol i Cd o the survivl d choliesterse ctivity of rigecked phest chicks d to compre the reltive cotriutios to toxicity of exposure through cotmited isect prey versus cotmited vegettio. METHODS Study site The field/pe study ws coducted withi -h psture of tive grssld elogig to the Brooks Horticulturl Reserch Sttio, Alert Miistry of Agriculture, Food d Rurl Developmet, locted km south of Brooks, Alert, Cd, i the mixed-grss pririe ecoregio. Vegettio ws predomitly lue grm grss (Boutelou grcilis) d sge (Artemesi c d A. frigid). The psture ws ugrzed d uured. Study imls Newly htched femle phest chicks were otied from the Brooks Wildlife Cetre (Alert Forestry, Lds, d Wildlife, Brooks, AB, Cd) d kept i idoor rooder pes i groups of 8 to for d. They were provided with cocetrted turkey strter rtio (Newlife Feeds, Lethridge, AB, Cd, 9% protei) d wter, d the temperture ws mitied t 5 to 8 C. At d old, chicks were rdomly ssiged to tretmet groups (six per group), weighed (. g) usig 5-g sprig scle (Pesol, Switzerld), mesured (trsus legth) usig Dil metric clipers (. mm), d mrked with umered squres of colored ylo tpe glued to fethers o their cks etwee the wigs. The comitio of tpe color d umer uiquely idetified ech ird. Experimetl procedure Betwee 5 d 7 h o dy, field plots were spryed usig icycle-mouted pressurized spryer desiged to deliver pesticide i the sme mer s typicl trctor-mouted spryers. Pressure ws 75 kp, d the spry volume ws L/ h. s icluded Furd 8F (crofur flowle, 8 g.i./l, Miles Ic., Mississug, ON, Cd), Cygo 8E (dimethote emulsifile, 8 g.i./l, Cymid Ltd., Mrkhm, ON, Cd), d Lors E (chlorpyrifos emulsifile, g.i./l, DowElco, Newmrket, ON, Cd). s were pplied t oe d four times the highest rtes recommeded for grsshopper cotrol i wester Cd [7]. These rtes were chose to represet wht ws ssumed to e typicl field scerios d to ecompss the possiility of ccidetl multiple-swth overlps, tk-mixig errors, d higher rtes used for other crops. Rtes were s follows: () Furd, d 58 g.i./h; () Cygo, d 85 g.i./h; d (c) Lors, 79 d, g.i./h. Formulted products were mixed with tp wter (ph 7.9) immeditely efore spry pplictio. Spry deposit ws mesured y plcig two rows of four filter ppers (9-mm dimeter) o the soil surfce t itervls cross the width of the swths t copy height (5 cm) for ech pplictio; filter ppers withi ech row were pooled for residue lysis. Ech spry swth ws m wide d m log, which ws sufficiet to ccommodte two pes ( m wide m log m high) seprted y m. Pes were pired i plots, with oe housig chicks fed ucotmited food d the other housig chicks fed grsshoppers tht hd ee spryed t the sme rte s the plot. This procedure llowed for compriso of reltive effects of cotmited food plus vegettio versus cotmited vegettio loe for ech isecticide t the two rtes of pplictio. Immeditely fter ll plots hd ee spryed, pes were erected over the spryed grss cover. Pes were costructed of.5-cm mesh chicke wire secured to metl corer supports covered with 5-cm mesh chicke wire. Ech ws equipped with woode rooder ox ( cm) plced i oe corer d heted with red icdescet -wtt lightul. Food dishes were plced o smll squres of plywood to reduce cotmitio from spryed vegettio d to ctch spilled food items i order to fcilitte mesuremet of food cosumptio. Dishes were kept t the etrce of the oxes durig the dy to keep food sheltered from wid d sulight. Wter dishes were plced outside er the rooder ox durig the dy ut were moved iside t ight. Dishes were checked periodiclly, d preweighed mouts of the ppropritely treted food ws dded s eeded to miti costt supply. Uete food ws removed d weighed. Totl food cosumptio per pe ws tllied over the 8-h exposure period, d the mout of food ete per chick-dy ws clculted for ech pe. Chicks were fed exclusive diet of grsshoppers throughout the 8-h exposure period. Three species of grsshoppers were collected from the field d spryed with field-equivlet rtes of the three isecticides. The species used were the cler-wiged grsshopper (Cmul pellucid), the two-striped grsshopper (Meloplus ivitttus), d the lesser migrtory grsshopper (Meloplus sguiipes). solutios were prepred volumetriclly with tp wter (ph 7.5) d spryed with lortory spry chmer (Iovtive Equipmet, Rogers Egieerig, Ssktoo, SK, Cd). The spry volume for ll isecticide tretmets ws equivlet to field pplictio volume of 8 L/h. The isecticides were spryed through sigle T-Jet SS8E ozzle (-mesh scree) t to kp. Amiet spry chmer temperture durig spryig ws to C. To miti equl volume of output mog tretmets, the selected ozzle trck velocity depeded o the isecticide product d cocetrtio d rged from. to.8 m/s. Spry droplet size d cocetrtio were moitored every fourth pss of the spry ozzle with wter-sesitive pper d with filter ppers for residue lysis. Droplets i six rdomly selected.5.5-cm res per tretmet were couted d mesured uder dissectig microscope to esure equl coverge for ll isecticide tretmets. Grsshoppers were lightly esthetized y directig strem of CO gs ito ech cotier for s. The resultig -mi period of esthesi provided eough time for spry pplictio without recovery d movemet of the isects. Aesthetized grsshoppers were rrged cm elow the ozzle d spryed oce. Grsshoppers i the cotrol groups were esthetized, hdled, d cged i the sme wy s the treted grsshoppers ut were ot spryed. d uspryed isects were froze d trsported to the field for feedig to chicks i the cges. The pplictio sequece ws replicted ccordig to the umer of cges to e fed. Overll there were tretmet groups, icludig cotrols (three isecticides, two pplictio rtes, d two food sources, i.e., spryed or uspryed). Three cotrol pes were icluded i ech replicte, d three complete replicte trils were coducted. Chicks were rought to the field fter ll the pes were erected (t to h) d were plced i the pproprite pes. Food ws provided immeditely. At pprox. h ech eveig, chicks were locked iside the rooder oxes to esure tht they styed wrm durig the ight. They were relesed d

3 5 Eviro. Toxicol. Chem. 5, 99 P.A. Mrti et l. give fresh food d wter etwee d 8 h oth susequet morigs. Birds were oserved from to 5 m wy t - to -h itervls durig dylight hours. Moriud irds were moitored closely util deth or recovery. If deth occurred, chicks were chilled o ice immeditely d tke to the lortory freezer ( C) withi h. Birds foud ded t the first morig check were chilled immeditely. Survivig chicks were scrificed usig cro mooxide poisoig t the ed of exposure. The ris of ll chicks were excised d froze i liquid itroge for susequet lysis of cetylcholiesterse (AChE) ctivity. The totl exposure period ws 8 h. Acetylcholiesterse lysis Alysis for AChE ctivity ws coducted t the Ntiol Wildlife Reserch Cetre (Hull, PQ, Cd). Bris were homogeized i chilled Tris uffer (ph 8.) usig glss tissue griders. Homogetes were ssyed colorimetriclly t m t 5 C usig cetylthiocholie iodide (Sigm Chemicl Co., St. Louis, MO, USA) s the ezyme sustrte [8,9]. Activity ws expressed s micromole sustrte hydrolyzed per miute per grm ri tissue (wet weight). residue lysis Alyses were coducted y EviroTest Lortories (Edmoto, AB, Cd). Filter-pper field smples were shke with ml of dichloromethe. Grsshoppers were extrcted twice y polytroig with : cetoitrile : hexe. Extrcts were comied d the cetoitrile dried through regulr sodium sulphte. The remiig hexe ws prtitioed with more cetoitrile, which ws the reduced to fil volume for lysis. Smple liquots were tke for isecticide lysis y gs chromtogrphy/mss spectrometry usig Hewlett-Pckrd (HP) 589 GC with HP 7 colum ( m.5 mm i.d.) coupled with HP 5975 mss selective detector. Detectio limits for filter-pper smples were, 5, d g for crofur, dimethote, d chlorpyrifos, respectively; for grsshoppers they were.,.5, d. g/g for the three isecticides, respectively. Recoveries for crofur, dimethote, d chlorpyrifos were 95, 7, d 7% from spiked filter-pper smples d, 7, d 97% from spiked grsshopper homogete. Reported vlues hve ee corrected for recovery efficiecy. Sttisticl lyses Sttisticl lyses were coducted usig the CATMOD d GLM procedures of SAS []. Differeces i mortlity mog the tretmet groups pooled cross ll replictes were tested usig G-tests. A iitil lysis of vrice (ANOVA) usig split split plot desig ws used to exmie the overll effects of isecticide, pplictio rte, d food cotmitio regime o dily growth rte (chge i weight d trsus legth) d ri AChE ctivity. ws the mi plot fctor; rte, the split plot fctor; d food, the split split plot fctor. To test the mi plot fctor, the isecticide replicte me squre ws used s the error term; to test the rte d rte isecticide fctors, the replicte rte isecticide me squre ws used s the error term. The effect of food d its iterctios were tested with the model error me squre. Cotrol groups were omitted from this lysis. Orthogol cotrsts were coducted to test priori questios of specific iterest []. We susequetly coducted ested ANOVA o ech pesticide seprtely, icludig pooled cotrol groups i the lyses for comprisos, usig the tretmet replicte me squre s the Tle.Spry deposit d chemicl residues i grsshoppers Lors Cygo Furd rte (g.i./ h) 79, Spry deposit (g.i./h) Me SE 9, 8, % 9 Coc. ( g/g) No. No Oe composite filter-pper (four per pool) smple per replicte tril. Pooled grsshopper smples from two trck spryer pplictios. error term i testig the effects of tretmet. Mortlities d survivors were lyzed seprtely. We determied differeces etwee tretmet d cotrol mes usig Tukey s hoestly sigifict differece tests (PROC GLM). RESULTS Spry deposit d grsshopper residues The me spry deposit over the three replicte trils ws slightly higher th % of the omil rte for ll pplictios (Tle ). However, deposits i the first two replictes were etwee d % of omil for most pplictios, while those i the third replicte rged from to % of omil. Residue cocetrtios showed good cosistecy etwee the two pooled grsshopper smples prepred i the trck spryer (Tle ). Mortlity Of 7 phest chicks moitored durig the experimet, totl of 7 died d 7 displyed symptoms of choliesterse ihiitio. The umer of chicks tht died or showed symptoms durig isecticide exposure did ot differ mog tretmet groups (Tle ). I replicte, lmost ll mortlities occurred durig the first ight; chicks were ded i the rooder oxes t the first morig check. Three chicks i the high-rte Lors group fed cotmited food died h fter the first morig check followig feedig out. of toxicity were ot oserved. I replicte, the oly two chicks tht died were i the low-rte Furd group receivig ucotmited food. Oe chick died out h fter iitil exposure d h fter relese from the rooder ox; the secod ird ws ded upo relese from the rooder ox o the secod morig. The third replicte hd the highest levels of mortlity overll; ll ut two deths occurred durig the secod ight of exposure, proly result of the uusully cold, dmp wether. The two erlier deths, oe i the low- rte Lors group receivig ucotmited food d oe i the low-rte Cygo group fed spryed food, occurred t out h exposure, to h fter relese from the rooder oxes. The chick i the Cygo group uderwet spsms d prlysis prior to deth. Growth d food cosumptio I the split split plot ANOVA, the mi fctors of isecticide, spry rte, or food cotmitio did ot ffect trsl growth or weight gi, lthough weight gi ws sigifictly ffected y the iterctio of the three fctors (p.5). Lrge differeces were oserved i the rtes of weight gi etwee irds tht survived d those tht died, pooled cross

4 Effects of isecticides o phests Eviro. Toxicol. Chem. 5, 99 5 Tle.Numer of phest chicks tht died or exhiited symptoms of choliesterse ihiitio durig 8 h of exposure to Lors, Cygo, or Furd i three replicte trils of tretmet regimes plus utreted cotrols rte (g.i./h) Food Replicte Ded Replicte Ded Replicte Ded Totl Ded Lors 79, 85 Cygo Furd 8 Cotrol Food regimes cosisted of either uspryed grsshoppers (uspryed) or grsshoppers spryed t the sme rte s the vegettio for tht tretmet (spryed). icluded lethrgy, persistet hed twitchig d gulpig, d spsms ll tretmet groups, icludig cotrols (Tle ) (p.); overll me chges i weight were.8 d. g/d, respectively. Rte of trsl growth showed similr differece (p.); overll gi i trsus legth ws. d. mm/ d for survivors d mortlities, respectively. Therefore, whe ested ANOVAs were coducted o idividul isecticides, we lyzed survivors d mortlities seprtely. Me rte of chge i ody weight of survivors of ll isecticide tretmets ws lower th tht of cotrols. Nevertheless, lthough the overll tretmet effect ws sigifict i the groups exposed to Cygo d Furd compred to cotrols (p.), the oly tretmet me tht differed sigifictly from tht of the cotrol group ws the high-rte Furd group cosumig spryed grsshoppers (.8 vs.. g/d, Tle ). Birds exposed to oth the high-rte Lors d the low-rte Cygo Tle.Me dily chges i ody weight (g) of phest chicks tht survived or died durig 8-h exposure to oe of three isecticides compred to utreted cotrols rte (g.i./ h) Lors 79, Cygo 85 Food Survivors Me SE Mortlities Me SE Furd Cotrol Food regimes cosisted of either uspryed grsshoppers (uspryed) or grsshoppers spryed t the sme rte s the vegettio for tht tretmet (spryed) Me sigifictly differet from cotrol me (ested ANOVAs withi isecticide, compred to pooled cotrols i ech lysis, Tukey s hoestly sigifict differece test, p.5. receivig uspryed grsshoppers hd me rtes of growth of less th 5% tht of cotrols (.9 d. vs.. g/d, respectively, Tle ), lthough differeces were ot sigifict. Rte of trsl growth i survivig chicks ws uffected y y of the isecticide tretmet regimes i compriso to cotrols (p.). Neither growth prmeter ws ffected y isecticide tretmets i chicks tht died durig exposure (p.). No sigifict differeces i food cosumptio were oserved mog y of the tretmet regimes d the cotrol group (Tle ). Bri cetylcholiesterse All three mi fctors i the split split plot ANOVA, isecticide, rte, d food cotmitio, sigifictly ffected AChE ctivity of phest chicks (p vlues.,.5, d., respectively). Overll me AChE ctivity of Furd-exposed groups ws less th the mes for groups ex- Tle.Food cosumed per chick per dy durig exposure to Lors Cygo, or Furd tretmets rte (g.i./h) Food Grsshoppers ete (g wet weight) Me SE Lors 79, Cygo Furd Cotrol Food regimes cosisted of either uspryed grsshoppers (uspryed) or grsshoppers spryed t the sme rte s the vegettio for tht tretmet (spryed) Me of three replicte trils. Food cosumptio ws mesured o per-pe sis, ot per-chick sis, ut corrected for umer of chicks per pe.

5 5 Eviro. Toxicol. Chem. 5, 99 P.A. Mrti et l. Tle 5.Bri cethylcholiesterse (AChE) ctivity (s % cotrol) of phest chicks either dyig durig or survivig 8-h exposure to oe of three isecticides rte (g.i./ h) Lors 79, Cygo 85 Furd 58 Food Survivors Me SE d d d 77. d. d Mortlities c Me SE Food regimes cosisted of either uspryed grsshoppers (uspryed) or grsshoppers spryed t the sme rte s the vegettio for tht tretmet (spryed). Me ri AChE ctivity of utreted (cotrol) survivors ( 8) ws.8 (SE.5) mol cetylthiocholie hydrolyzed/mi/g ri tissue. c Me ri AChE ctivity of utreted (cotrol) mortlities ( ) ws. (SE 5.5) mol cetylthiocholie hydrolyzed/mi/g ri tissue. d Mes sigifictly differet from cotrol group me (ested AN- OVAs withi isecticide, compred to pooled cotrols i ech lysis, Tukey s hoestly sigifict differece test, p.5). posed to Cygo or Lors; me AChE ctivity of irds exposed to high spry rtes ws lower th tht of those exposed to low spry rtes; d irds cosumig spryed food hd lower AChE ctivity th those fed uspryed food. A pled orthogol cotrst showed overll me ri AChE ctivity of irds i the Furd tretmets ws slightly lower th tht of irds i ll Cygo d Lors tretmets together (p., mes of 9. vs..5 mol/mi/g). Overll, irds tht died hd sigifictly higher ri AChE ctivity th those tht survived (.9 vs..5 mol/mi/g, p.), so further lysis y pesticide ws coducted o these two groups seprtely. Tretmet hd sigifict effect i irds survivig the Cygo d Furd regimes (p.). Bri AChE ctivity ws lower i irds survivig exposure to Cygospryed grsshoppers t oth low d high rtes th i cotrols (Tle 5). Birds fed Furd-treted grsshoppers t oth spry rtes, s well s those exposed to the high-rte vegettio cosumig utreted food, exhiited sigifictly depressed ri AChE ctivity (Tle 5). Me ri AChE ctivity of irds fed uspryed grsshoppers d exposed to vegettio treted with high-rte Furd ws lower th tht of irds etig food treted t the lower rte. Bri AChE ctivity of irds tht died durig exposure to y of the isecticide regimes did ot differ sigifictly from tht of cotrols (Tle 5). DISCUSSION Depositios of the three grsshopper-cotrol isecticides mesured i our spryed plots were somewht higher th those detected i eril pplictios reported to hve rged from 8 to % of trget rte [ ]. This vritio i pesticide depositio provides iformtio for the worst-cse scerio t give spry rte. Grsshopper cocetrtios were cosistet etwee the two spryig evets used i their preprtio. I previous study, grsshoppers collected followig eril pplictios of Furd t g.i./h to two pstures cotied me crofur cocetrtios of. d. g/g wet weight for live grsshoppers d.9 g/g for ded grsshoppers [5]. Grsshoppers i the gullets of Clifori gulls tht died o griculturl ld recetly spryed with Furd cotied. to 7. g/g crofur [5]. Thus, cocetrtios i our lortory-spryed grsshoppers pper to e represettive of cocetrtios i isects ville to irds forgig i the field. Despite the low levels of mortlity, sigs of ticholiesterse toxicity were oserved i 7 of pesticide-exposed phest chicks (7.9%); six of these occurred i the pe treted with the high-rte Furd d receivig spryed food. Most irds recovered withi few hours of the oset of symptoms d ppered to suffer o prologed effects. Nevertheless, this period of sickess my hve serious cosequeces for roods i the wild. Chicks of precocil species requirig pretl roodig d directio my e doed y their prets durig spell of icpcittio, therey reducig their chces of survivl. Mrti d Forsyth [8] foud tht mllrd hes movig overld with their roods would wit oly riefly (miutes) for Furd-itoxicted ducklig d would soo doed it. Treds of reduced weight gi lso were oserved i irds exposed to isecticide-spryed grss, prticulrly grss treted t the high spry rtes, suggestig some loger-term impcts o phest chick growth. I prticulr, irds exposed to the grss treted t the high spry rte of Furd d fed spryed grsshoppers grew more slowly th cotrol irds. However, 8-week-old turkeys livig i rge pes where grss d soil hd ee spryed with much higher levels of chlorpyrifos th used i the preset study ( d kg.i./h) for 8 d chieved the sme ody weight s cotrol irds [W.S. McGregor d R.W. Swrt, upulished dt]. All chicks tht died durig the exposure period, icludig cotrols, gied weight sigifictly more slowly th survivig chicks, d most ctully lost weight. -exposed irds experieced o greter loss i weight th did cotrol irds, however, idictig miiml dditiol impct of isecticides s fctor cotriutig to weight loss. Bri AChE ctivity ws ot depressed i exposed chicks tht died, further suggestig isigifict role plyed y cute isecticide toxicity i this study. Some irds were foud ded t the first check i the morig; thus, time of mortlity ws ukow. It is possile, therefore, tht some postmortem rectivtio of ri AChE ctivity my hve occurred. Bri AChE ctivity of Jpese quil dmiistered sulethl dose of crofur ( mg/kg) d euthized h lter hd rectivted to cotrol levels h postmortem whe held t 5 C []. I cotrst, similr dose of the orgophosphorus compoud dicrotophos cused cotiued declie i AChE ctivity for 7 h postmortem []. At lethl dosges of crofur, however, despite slight postmortem rectivtio, ri AChE ctivity ws still sigifictly lower th i cotrols 7 h fter deth; ctivity of dicrotophos-dosed irds cotiued to declie durig this period []. Bri AChE ctivity ws eqully depressed immeditely d 7 d postmortem i Jpese quil (Coturix jpoic) lethlly dosed with dimethote [7]. If ri AChE ctivity of phest chicks i the preset study hd ee depressed to the levels usully ssocited with deth ( 5% of cotrol ctivity [8]), sigifict depressio should still hve ee detectle despite the dely i ri removl d cold-storge fter deth. It is likely, therefore, tht

6 Effects of isecticides o phests Eviro. Toxicol. Chem. 5, 99 5 these irds died ecuse of strvtio or hypothermi excerted y the stresses of the outdoor peig situtio despite the provisio of rooder oxes, which would expli the deths i cotrol irds s well. Although ticholiesterse isecticides hve ee show to reduce thermoregultory cpility i the youg of precocil species [5,9], this ws proly ot fctor i the deths of our isecticide-exposed chicks ecuse they suffered o greter mortlity th did cotrol chicks. Sigifict reductios i the thermoregultory ilities of youg duckligs occurred oly whe ri AChE ctivity ws ihiited y greter th 5% [5]. Cosiderle vriility i the rtes of weight gi mog irds withi tretmet groups ws oserved. This vriility my hve idicted greter sesitivity of some idividuls to the tste or toxic effects of the isecticide. Some irds my hve voided the food ecuse of either tste versio or coditioed versio cused y ssocitig food with ticholiesterse symptoms [,]. Other idividuls withi group must hve ee le to cosume dequte qutities of food to miti orml growth rtes while voidig lethl dose. This is further evideced y the fct tht, o pe sis, chicks i tretmet groups receivig isecticide-cotmited food sources did ot cosume less food th cotrols. Hlf of group of orther owhite (Colius virgiius) chicks exposed to food cotiig crofur t 59 g/g for 5 d reduced their cosumptio of treted d cle food comied y 5% reltive to cotrol irds []. Phest hes reduced cosumptio y 8% whe provided with Furd-treted food (crofur t 5 g/g) d y 9% whe provided with Lors-treted food (chlorpyrifos t, g/g) loe []. Although food cosumptio ws ot mesured, -d-old phests fed Furd-cotmited gri s their sole food source (crofur t.55 g/g) grew t the sme rte s cotrol chicks []. Clycolored sprrows fed grsshoppers cotiig crofur t. g/g cosumed more food th did those provided with uspryed grsshoppers d showed o toxic effects or y loss i weight [5]. Although the phest chicks i our study tht lost weight d died my hve ee those tht were voidig food, it is lso prole tht, s similr umers of deths d equl rtes of weight loss occurred i the cotrol groups, other fctors were resposile. For exmple, domit phest chicks my prevet iferior chicks from ccessig the food dish, resultig i differetil weight chges d survivl []. The cute orl medi lethl doses (LD5s) of chlorpyrifos, dimethote, d crofur for -d-old phest chicks dosed usig topicl pplictio to grsshopper crcsses re 5.8, 8.9, d.77 mg/kg ody weight [P.A. Mrti d D.L. Johso, upulished dt]. Give typicl ody weight of g for -dold phests, chicks i our study were cosumig mximum doses equivlet to.,., d 8.5 LD5 doses per dy for the three isecticides, respectively (Tle ). The lck of toxic effects oserved i our study my e due to the ptter of isecticide itke, distriuted over pprox. h of dylight rther th s cute dose. Crofur i prticulr is very quickly metolized due to the rpid decrmyltio of AChE [7]. Depressed AChE ctivity of chicks exposed to cotmited food i the Furd d Cygo tretmets idicted tht irds were receivig their primry itke of the isecticides i their diet rther th from the spryed vegettio. Nevertheless, AChE ctivity ws lso reduced i chicks i the high-spryrte Furd tretmet receivig ucotmited food, lthough depressio i this group ws cosiderly less th tht i the similr regime receivig cotmited food ( vs. %). Tht Lors Cygo Furd Tle.Pesticide itke y phest chicks feedig o grsshoppers cotmited with oe of three isecticides rte (g.i./h) 79, coc. ( g/g) Food cosumed (g/chickdy) Dily dose ( g/g ody wt) Dose/ LD Clculted (food cocetrtio [ g/g] food cosumed [g])/ g, verge ody weight of -d-old phest chick. Bsed o vlues from Mrti d Johso (upulished dt) for - d-old phest chicks. there ws sigifict depressio i the uspryed food tretmet suggests tht phest chicks were proly forgig for turl food i the spryed grss d possily receivig isecticides y preeig cotmited fethers. Driver et l. [8] determied tht preeig d igestio of cotmited food cotriuted eqully to the depressio of ri AChE ctivity i dult orther owhites exposed to simulted crop hitt spryed with methyl prthio t. kg/h i wid tuel. Quil i tht study were preset i the crop durig spryig, however, which my hve icresed the mout of isecticide o the irds fethers s well s uptke through ihltio. Derml uptke through the feet is other method tht my hve cotriuted to the depressed AChE ctivity of phest chicks i our study [8]. Tht depressio did ot occur i chicks provided with uspryed food t the lower Furd rte or either of the two orgophosphorus isecticide rtes i our study suggests tht uptke of isecticides vi preeig, forgig i spryed grss, or derml exposure is isufficiet to ffect chicks uless very high levels of very potet choliesterse ihiitor re used. These fidigs corroorte those of Mrti et l. [7] d Mrti d Forsyth [8] tht mllrd duckligs exposed to upld vegettio spryed t oe d two times the recommeded grsshopper- cotrol rtes of Furd hd to cosume cotmited food items from the vegettio i order to oti sufficiet isecticide to hve sigifict impct o choliesterse d ehvior. The lck of sigifict mortlity of phest chicks i grssld plots freshly spryed with isecticides t oe d four times the rtes of pplictio recommeded for grsshopper cotrol suggests tht the risk of cute toxicity preseted y these chemicls to juveile glliform species is firly low. Behviorl effects ppret i the smll umer of chicks udergoig sulethl itoxictio i our study could result i decresed survivl of ffected idividuls i free-livig roods. As well, the tedecy towrd lower weight gis i chicks i spryed plots might e excerted i the wild y reductios i rthropod food resources throughout rood s forgig rge. Evidece from documeted ird kills [,5] suggests tht dults of species tht gorge themselves rpidly i spryed res, such s wterfowl d gulls, my e more vulerle to cute isecticide poisoig compred to juveile irds tht cosume much smller qutities of cotmited food over prologed period of time. AckowledgemetThe dvice d logistic support of D. Jcoso is much pprecited. C. Adrews provided ssistce i the field d with the rerig d spryig of grsshoppers.

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