S. N. Thompson*, R. A. Redak and L.-W. Wang Department of Entomology, University of California, Riverside, California 92521, USA

Transcription

1 The Journl of Experimentl Biology 28, Published by The Compny of Biologists 25 doi:1.1242/jeb Host nutrition determines blood nutrient composition nd medites prsite developmentl success: Mnduc sext L. prsitized by Cotesi congregt (Sy) S. N. Thompson*, R.. Redk nd L.-W. Wng Deprtment of Entomology, University of Cliforni, Riverside, Cliforni 92521, US *uthor for correspondence (e-mil: ccepted 24 November 24 This investigtion exmined the influence of dietry protein nd crbohydrte blnce in chemiclly defined rtificil diet for Mnduc sext lrve on development of the gregrious prsite Cotesi congregt. Norml unprsitized lrve nd lrve superprsitized in the fourth stdium were rered to the end of the fifth stdium on six diets, ech hving the sme totl mount of csein nd sucrose but with different rtios rnging from high protein/no crbohydrte through to low protein/high crbohydrte. Levels of blood protein nitrogen nd trehlose, nutrients supporting growth nd development of C. congregt, vried with diet nd were influenced by prsitism. Different levels of blood metbolites reflected differences in diet consumption, nd the reltionships between protein nitrogen nd trehlose were very similr to those for protein nd crbohydrte intke by prsitized nd norml lrve on vrious diets. Dietry nutrient rtio hd significnt effect on prsite burden, the numbers of prsites developing in individul host lrve nd on prsite biomss. Prsites included individuls tht developed nd eventully emerged s second instr lrve, moulted to third instrs nd pupted. Mny pprently mture second instr prsites, however, filed to emerge. The proportion of nonemerging individuls vried with diet, nd in some cses, prsites filing to emerge were greter in number nd Summry totl biomss thn those tht did emerge to complete development. On most diets, the mss of individul prsites ws similr regrdless of dietry nutrient rtio. Three dimensionl models developed to demonstrte the reltionships between blood protein nitrogen nd trehlose levels nd prsite burden nd biomss estblished tht the levels of both metbolites re importnt for supporting growth nd development of emerged nd non-emerged prsites. In the cse of emerged prsites, however, the reltionships re liner, nd qudrtic function best describes the reltionships with non-emerged prsites. Blood metbolite levels supporting the gretest prsite burden nd biomss of emerged nd non-emerged prsites occupy region of two dimensionl spce corresponding to pproximtely 6 2 mg per insect of protein nitrogen nd 6-1 mg per insect of trehlose. Despite the differences in the response of emerged nd non-emerged prsites to host nutrition, the present results indicte tht host nutrition is not the criticl fctor determining prsite emergence. The significnce of these findings to the biology of C. congregt is discussed. Key words: insect, development, nutrition, diet, prsitism, Mnduc sext, Cotesi congregt. Introduction Prsitism is the ssocition of two orgnisms where one, the prsite, lives t the expense of the other, the host (Roberts nd Jnovy, 2). Prsites re hrmful nd in mny cses the host ultimtely dies (Ewld, 1995). Nevertheless, host survivl if only temporry is essentil for the success of most prsites (Smyth, 1976; Thompson, 1985). Nourishment nd development of hymenoptern insect endoprsites is highly integrted with the biology of their insect hosts (Mckuer nd Sequeir, 1993; Quickie, 1997). Prsitized hosts undergo complex physiologicl ltertions tht ensure suitble conditions for prsite development (Vinson nd Iwntsch, 198; Thompson, 1993; Beckge nd Gelmn, 21). During the ssocitions of mny insect endoprsites, including gregrious brconid wsps of the genus Cotesi, with their lepidoptern lrvl hosts, the effects of prsitism re initilly medited by vriety of prsite-derived fctors. These include polydnviruses nd/or venom, injected into the host by the dult femle prsitoid during oviposition (Beckge et l., 1994; Nkmtsu et l., 21; Nkmtsu nd Tnk, 23). lso involved re tertocytes, specilized cells derived from the serosl membrne of the prsite egg (Dhlmn nd Vinson, 1993; Zhng et l., 1997). criticl role of such components is erly suppression of host defense responses tht would otherwise encpsulte nd destroy the prsite egg or erly lrvl stges (Schmidt et l., 2). n dult femle Cotesi spp. deposits between 5 nd few hundred eggs into

2 626 S. N. Thompson, R.. Redk nd L.-W. Wng the body cvity or hemocoel of single host lrv. Host lrve, however, re often superprsitized, where more thn one dult prsitoid oviposit eggs. The eggs htch nd prsite lrve develop, feeding principlly on the host s hemolymph or blood, but lso on the ft body following disintegrtion through the ction of tertocyctes (Nkmtsu et l., 22). In the cse of Mnduc sext prsitized by Cotesi congregt, mture, second instr prsite lrve emerge from the host, moulting to the third stdium s they penetrte the cuticle (Fulton, 194). Upon moulting, prsite lrve spin cocoons nd pupte. Prsitism lso brings bout long-term physiologicl effects, mny of which my influence the ultimte success of prsite development. Depressed host growth nd incresed development time re common responses (Vinson nd Iwntsch, 198; Beckge nd Riddiford, 1983). Delyed host development my be importnt for ensuring sufficient time for prsite growth nd development (Smith nd Smilowitz, 1976; Slnsky, 1978; Lwrence nd Lnzrein, 1993). Numerous studies, including investigtions of vriety of lepidoptern insects prsitized by Cotesi spp. demonstrte tht decresed food consumption ccompnies the bove effects (Tnk et l., 1992; lleyne nd Beckge, 1997). Otherwise, little is understood of the potentil effects of nutrition on prsitized host insects or of the importnce of host nutrition to prsite success. Recent studies estblish tht dietry nutrient blnce influences growth nd development of Mnduc sext over the lst two lrvl stdi (Thompson et l., 25). Feeding nd nutrient intke differ mrkedly between norml nd prsitized lrve nd depend on the rtio of digestible protein nd crbohydrte. Growth of prsitized lrve is equivlent to tht of norml unprsitized lrve when insects re mintined on diets hving rtio of csein (C) to sucrose (S) between 1.C:1.S nd 1.5C:.5S, lthough development time of prsitized lrve is longer. On diets hving nutrient rtios with greter or lesser protein, growth of prsitized lrve is severely depressed when compred with norml lrve even though development times of prsitized insects re greter on the less suitble diets. These results suggest tht prsitized lrve my exhibit different long-term feeding preferences thn norml lrve. Tht conclusion is consistent with results of previous investigtion demonstrting tht prsitized lrve, offered choice of diets hving vrible nutrient content, select different rtio of nutrients from tht preferred by norml lrve (Thompson et l., 21). Understnding how host responses to nutritionl vrition medite prsite growth nd development is importnt to ssess whether nutritionl fctors ply criticl role in successful prsitism. The present study ddresses how dietry protein nd crbohydrte blnce influences the development of C. congregt in M. sext. We estblish how host dietry nutrient rtio ffects the totl levels of blood protein nitrogen nd trehlose, nd in turn, how these metbolites ffect prsite burden, the numbers of prsite lrve developing in individul hosts nd totl prsite biomss. Furthermore, we exmine the reltionship between host dietry nutrient rtio, blood metbolites nd the numbers of mture prsite lrve tht emerge from the host to pupte nd complete development nd the numbers of lrve tht fil to emerge. Bsed on the findings of studies cited bove, we predicted tht the nutritionl sttus of host lrve s ffected by dietry nutrient rtio would influence prsite burden nd success through effects on host blood composition. Mterils nd methods Insect culture Stock colonies of Mnduc sext L. were rered on semidefined whet germ-bsed rtificil diet s previously outlined (Thompson et l., 25). Lrve used in the experiments were rered on the rering diet until completion of the third stdium. Phrte fourth instr host lrve were synchronized s described by others (Bker et l., 1987) nd superprsitized. For prsitiztion, lrve were plced individully in 3 l glss jrs contining 1 15 Cotesi congregt (Sy) prsitoids of both sexes. Lrve were prsitized two to four times (lleyne, 1995), removed from the jrs nd plced on the experimentl diets in 16 ml plstic cups. Experimentl diet nd feeding protocol Newly moulted fourth instr M. sext lrve were fed chemiclly defined rtificil diet contining vrible levels of csein nd sucrose s digestible protein nd crbohydrte, respectively (hmd et l., 1989). The diet lso contined B vitmins, linseed oil, scorbic cid nd Wesson s slt mixture, obtined principlly from Bioserve (Frenchtown, NJ, US) nd Nutritionl Biochemicls (Clevelnd, OH, US). Six diets were employed, ech hving the sme totl mount of csein nd sucrose, but with different csein to sucrose rtios s follows:.125c:1.875s,.25c:1.75s,.5c:1.5s, 1.C:1.S, 1.5C:.5S nd 2.C:S. The nutrient levels re indicted reltive to the mount of csein nd sucrose in the stock formultion, tht is, 1C:1S reltive to 9 g l 1 csein nd 9 g l 1 sucrose. Groups of 1 rndomly selected norml nd prsitized lrve were mintined on ech experimentl diet for the feeding studies described below. Insects were housed in Precision Scientific incubtor t 28 C with 16 h:8 h light:drk non-dipusing, long-dy photocycle. Lrve were fed on the experimentl diets from the strt of the fourth stdium until the end of the fifth stdium. In the cse of norml lrve, the experiments were discontinued fter pproximtely 25% of the lrve hd stopped feeding nd entered the wndering phse in preprtion for puption. t this point ll lrve hd reched pproximtely 8 1 g with totl development time between 7 nd 15 dys, depending upon diet. For prsitized lrve, the experiment ws stopped t the time lrve cesed feeding prior to prsitoid emergence. The time lso vried with diet, between 12 nd 15 dys. Estimtion of prsite burden nd biomss The number of prsites tht emerged from individul host

3 Host nutrition medites prsite development 627 lrve mintined on the vrious diets ws determined t the end of the experiments by counting prsite lrve nd cocoons. fter dissection of the gut of prsitized lrve, the prsites filing to emerge were counted using Wild dissecting microscope. To determine prsite biomss, the emerged nd non-emerged prsites were collected nd dried in n oven t 1 C for 24 h. Biomss ws mesured on Srtorius microblnce. Estimtion of host blood metbolite levels Previously, we reported the effects of dietry nutrient rtio nd prsitism on equilibrium blood concentrtions (mg ml 1 ) of protein, totl free mino cids nd trehlose in norml nd prsitized M. sext lrve (Thompson et l., 25). Here, we use those dt to estimte the totl quntities or levels of these metbolites in the blood of norml nd prsitized lrve bsed on their finl mss nd wter content, ssuming 5% extrcellulr wter (Chpmn, 1998). Dt re presented s totl protein nitrogen level (protein plus free mino cids) nd totl trehlose (mg per insect). Determintion of host finl mss nd nutrient consumption Diet consumption ws determined s the difference between the totl mount of diet offered to lrve nd the mount remining in the diet cups t the end of the experiment together with undigested diet remining in the gut. Dry mss of the diet remining in the cups ws determined by drying the diet in n oven s described bove. Initil dry mss of the diet offered to the insects ws estimted from the known rtio of wet/dry mss. Protein nd crbohydrte consumption were estimted bsed on the composition of ech diet. The guts of norml nd prsitized lrve were dissected t the end of the experiment nd the diet remining in the gut ws removed. This diet ws dded to tht remining in the cups, for estimtion of diet consumption. The individul crcsses of host lrve were dried s bove nd finl host mss mesured by weighing. Generl sttisticl nlyses Dt showing the effects of dietry nutrient rtio on prsite burden nd biomss were exmined by two-wy nlysis of vrince (NOV). nlysis of covrince (NCOV) using vrious prmeters s covrites, were pplied in specific cses described below. The Shpiro-Wilk W test nd norml probbility plots evluted normlity nd homogeneity of vrince. Except in the cse of prsite burden, dt met the ssumptions for nlysis of vrince. Non-normlly distributed dt for prsite burden were squre root trnsformed. Results Effect of dietry nutrient rtio nd prsitism on host blood metbolite levels Dietry nutrient rtio nd prsitism ech ffected blood protein nitrogen level of M. sext lrve nd there ws no Tble 1. NOV summry demonstrting the effects of dietry nutrient rtio nd prsitism by Cotesi congregt on the totl levels (mg per insect) of blood protein nitrogen nd trehlose in fifth instr Mnduc sext lrve rered on chemiclly defined rtificil diet Men F Dependent vrible d.f. squre vlue Probbility Protein nitrogen (mg per insect) Dietry nutrient rtio <.1 Prsitism =.9 Interction =.2349 Error Trehlose (mg per insect) Dietry nutrient rtio <.1 Prsitism =.548 Interction =.4 Error interction between dietry nutrient rtio nd prsitism (Tble 1). Trehlose level ws lso ffected by dietry nutrients, but ny effect of prsitism ws mrginl. Further, there ws significnt interction between prsitism nd dietry nutrient rtio demonstrting tht dietry nutrient rtio ffects trehlose differently in norml nd prsitized lrve (Tble 1). The reltionship between blood protein nitrogen nd trehlose levels (mg/insect) reltive to nutrient consumption by hosts on the vrious diets is shown in Fig. 1. For norml Totl blood trehlose (mg/insect) C:1.75S.25C:1.75S.125C:1.875S.5C:1.5S 1.C:1.S 1.C:1.S 2.C:S 1.5C:.5S 5 1 Totl blood protein nitrogen (mg/insect) 1.5C:.5S 2.C:S Fig. 1. Two-dimensionl representtion of blood protein nitrogen (protein nd free mino cids) nd trehlose levels (mg per insect) in norml (filled circles) fifth instr M. sext lrve nd lrve prsitized by C. congregt (open circles), mintined over the fourth nd fifth stdi on chemiclly defined rtificil diet hving vrying rtios of csein nd sucrose. Vlues re mens ± S.E.M. Dietry nutrient rtios shown reltive to the level of ech nutrient (C, csein; S, sucrose) in the bsl chemiclly defined formultion where 1.=9 g l 1.

4 628 S. N. Thompson, R.. Redk nd L.-W. Wng unprsitized lrve, totl blood trehlose decresed s the dietry level of sucrose decresed, with the exception of the.125c:1.875s diet. This diet ws previously judged pthologicl (Rubenheimer nd Simpson, 1999) bsed on erlier findings tht norml host lrve fil to djust consumption in response to the poor nutrient blnce of this diet (Thompson et l., 25). In contrst, with prsitized lrve totl blood trehlose incresed s dietry level of sucrose decresed reching mximum on the 1.C:1.S diet, therefter decresing with further decreses in dietry crbohydrte. Totl blood nitrogen for both prsitized nd norml lrve incresed s dietry protein level incresed nd ws mximl on the 1.5C:.5S nd 2.C:.5S diets. Effect of host dietry nutrient rtio on prsite burden nd biomss Dietry nutrient rtio hd significnt effect on prsite burden, the numbers of prsites developing in individul host lrve nd totl biomss, regrdless of whether prsites emerged to complete development (Tble 2; Fig. 2). The prsite burden nd biomss of non-emerged prsites were gretest on the 1.C:1.S nd the 1.5C:.5S diets (Fig. 2B). Non-emerged prsite burden nd biomss were lower in host lrve on the other diets, but there ws no significnt difference between these diets. The burden nd totl biomss of emerged prsites ws more uniform between diets nd highest from host lrve on the.5c:1.5s, 1.C:1.S nd 1.5C:.5S diets (Fig. 2B). No prsites emerged from lrve on the.125c:1.875s diet. On this diet prsite burden ws very low nd the individul prsites were very smll (prsite prmeters were not determined). The mss of individul prsite lrve tht emerged or tht filed to emerge ws similr for host lrve t ll dietry nutrient rtios, except for Fig. 2. Effects of host dietry nutrient rtio on prsite burden (), totl prsite biomss (B) nd individul prsite mss (C) of C. congregt, emerged nd nonemerged from M. sext lrve mintined over the fourth nd fifth stdi on chemiclly defined rtificil diet hving vrying rtios of csein nd sucrose. Dietry nutrient rtios shown reltive to the level of ech nutrient (C, csein; S, sucrose) in the bsl chemiclly defined formultion where 1.=9 g l. Significnt differences (P<.5) between emerged prsites on different diets re indicted by different lower cse letters nd differences between nonemerged prsites by different upper cse letters. prsite burden Totl prsite biomss (g) verge individul prsite mss (mg) C B Emerged Non-emerged b c b B,C C B,b B,C B,C.25C:1.75S.5C:1.5S 1.C:1.S 1.5C:.5S 2.C:S Dietry nutrient rtio,b,b,b,b b,c C C

5 Host nutrition medites prsite development 629 Tble 2. NOV summry demonstrting the effects of host dietry nutrient rtio on Cotesi congregt burden nd biomss emerged nd non-emerged from fifth instr Mnduc sext lrve mintined on chemiclly defined rtificil diet Dependent vrible d.f. Men squre F vlue Probbility Prsite burden emerged Dietry nutrient rtio =.5 Error Prsite burden non-emerged Dietry nutrient rtio <.1 Error Totl prsite biomss (g) emerged Dietry nutrient rtio =.8 Error Totl prsite biomss (g) non-emerged Dietry nutrient rtio <.1 Error Individul prsite mss (mg) emerged Dietry nutrient rtio <.1 Error Individul prsite mss (mg) non-emerged Dietry nutrient rtio <.1 Error those from hosts on the.25c:1.75s diet (Fig. 2C), where both emerged nd non-emerged prsites weighed less. lrge portion of non-emerged prsites pper to be mture second instr lrve, n observtion previously reported by lleyne nd Beckge (1997). We compred the burden nd the biomss of emerged nd non-emerged prsites from hosts on the individul diets by conducting n NOV nlysis on the rithmetic differences between the two prsite popultions. The results (not shown) demonstrte significnt effect of dietry nutrient rtio on the men difference scores for both burden nd biomss. Only in the cse of the 1.C:1.S diet, however, were the difference scores between non-emerged nd emerged prsite burden nd biomss significntly different from, with emerged prsites being fewer nd hving less totl biomss. Effect of host blood nutrient levels on prsite burden nd biomss We estimted the totl levels of protein nitrogen nd trehlose in the insects for which prsite dt were vilble bsed on the totl metbolite levels reported bove. First, we estblished the reltionships between metbolite levels nd host finl mss by pplying NCOV using blood protein nitrogen nd trehlose levels s dependent vribles, dietry nutrient rtio s min-effect tretment nd finl mss s the covrite. Dietry nutrient rtio hd significnt effect on both metbolites nd in ech cse, there ws significnt covrite effect of finl mss (Tble 3). Using the metbolite levels predicted from liner regression equtions, we modeled the effect of protein nitrogen nd trehlose on emerged nd nonemerged prsite burden nd biomss (PROC REG followed by PROC G3 GRID nd PROC G3D. SS version Tble 3. NCOV summry demonstrting the effects of dietry nutrient rtio on protein nitrogen nd trehlose levels (mg per insect) in fifth instr Mnduc sext lrve mintined over the fourth nd fifth stdi on chemiclly defined rtificil diet hving vrying rtios of csein nd sucrose Finl mss (g) ws the covrite. Dependent vrible d.f. Men squre F vlue Probbility Protein nitrogen (mg per insect) Dietry rtio <.1 Finl mss (covrite) <.1 Error Trehlose (mg per insect) Dietry rtio <.1 Finl mss (covrite) =.8 Error

6 63 S. N. Thompson, R.. Redk nd L.-W. Wng SS Institute Inc., Cry, NC, US). Dt for protein nitrogen nd trehlose levels were first stndrdized (men of. nd stndrd devition of 1.) nd then used s dependent model vribles. Stndrd vrible selection techniques (Freund nd Little, 2) were used to generte best-fit model tht considered both liner nd qudrtic terms s independent vribles nd prsite burden or biomss s dependent vribles. Four predictive models were constructed. dditionlly, contour mps were generted from these dt (PROC GCONTOUR) from point mtrix by interpolting simple liner function for the reltionship between blood metbolite levels nd finl mss (PROC G3 GRID). liner model provided the best fit for emerged prsite burden nd biomss, nd qudrtic nd interction terms involving protein nitrogen nd trehlose were not included in these models (Tble 4). Blood protein nitrogen nd trehlose levels both predicted emerged prsite biomss (Fig. 3). The reltionship for prsite burden is not shown. Both protein nitrogen nd trehlose levels were positively ssocited with incresed emerged prsite burden nd biomss (Tble 5). Bsed on the stndrd regression coefficient, trehlose ws the more importnt predictor of prsite biomss, while protein nitrogen ws the more importnt in the cse of prsite burden. With non-emerged prsite burden nd biomss (Fig. 3C), both trehlose nd protein nitrogen levels were importnt, but the reltionships were more complex thn with emerged prsite prmeters (Fig. 3). Here, the qudrtic terms for both protein nitrogen nd trehlose significntly contributed to the precision of the models in predicting non-emerged prsite burden nd biomss (Tble 5). The model component describing the interction between trehlose nd nitrogen did not contribute to the bility of the models to predict non-emerged prsite burden nd biomss. Trehlose level (both liner nd qudrtic estimtes) ws the most importnt predictor of nonemerged prsite prmeters. However, the overll effect of trehlose ws similr to tht of nitrogen. t intermedite levels of both metbolites, fewer numbers of non-emerged prsites re produced. Low nd high levels of metbolites led to greter numbers nd biomss (Fig. 3C) of prsites filing to emerge. The contour mps illustrte the optiml levels of protein nitrogen nd trehlose supporting prsite growth nd development. The reltionships between blood metbolite levels nd emerged nd non-emerged prsite biomss re shown in Fig. 3C,D. Reltionships for prsite burden were similr but re not shown. Nutrient levels for the.5c:1.5s, 1.C:1.S nd 1.5C:.5S diets supporting the gretest numbers nd biomss of prsites pper to occupy common region in two dimensionl spce. Within this spce, the nutrient levels vried between pproximtely 6 nd 11 mg per insect Tble 4. NOV summry for models estimting the reltionship between blood protein nitrogen nd trehlose levels (mg per insect) nd emerged prsite burden nd biomss Prsite burden Source d.f. Men squre F vlue Probbility r 2 Model Error Model prmeter estimtes Stndrdized Stndrd regression error of Vrible coefficient coefficient Trehlose Nitrogen Intercept Prsite biomss Source d.f. Men squre F vlue Probbility r 2 Model Error Model prmeter estimtes Stndrdized Stndrd regression error of Vrible coefficient coefficient Trehlose Nitrogen Intercept

7 Host nutrition medites prsite development 631 Emerged prsite stndrdized biomss (mg) Non-emerged prsite stndrdized biomss (mg) C Blood trehlose (mg/insect) Blood trehlose (mg/insect) Blood protein nitrogen (mg/insect) Blood protein nitrogen (mg/insect) Blood trehlose (mg/insect) B D C:1.75S.5C:1.5S 1.C:1.S 1.5C:.5S 2.C:S Blood protein nitrogen (mg/insect) c:1.75s.5c:1.5s 1.C:1.S 1.5C:.5S 2.C:S Fig. 3. Multidimensionl profiles illustrting the effects of blood protein nitrogen trehlose levels (mg per insect) on biomss of C. congregt, emerged nd non-emerged from M. sext lrve mintined over the fourth nd fifth stdi on chemiclly defined rtificil diet hving vrying rtios of csein (C) nd sucrose (S). (,C) Three-dimensionl models for emerged nd non-emerged prsite biomss, respectively. See text for model specifictions nd detils. (B,D) Contour mps show ctul emerged nd non-emerged biomss, respectively. Blood metbolite levels re indicted for individul lrve on the vrious diets. trehlose nd between 6 nd 2 mg per insect protein nitrogen. Discussion The present study demonstrtes tht development of C. congregt depends on host nutrition, specificlly the effects of nutrient intke on the levels of host blood metbolites. s described bove, prsite lrve feed principlly on blood with blood metbolites providing nourishment for prsite growth nd development. lthough Cotesi spp. re thought to consume nutrients by orl ingestion, possibly ided by stomodel pump (Quickie, 1997; Nkmtsu nd Tnk, 22), first nd prticulrly second instr C. congregt lrve hve both unsclerotized cuticle nd well developed nl vesicles (Fulton, 194). The lter form by eversion of the rectum such tht the bsorptive epithelium is in direct contct with the externl medium. Studies with nother brconid species, Microplitis croceipes, hve demonstrted tht the nl vesicle ctively bsorbs mino cids nd sugrs, including trehlose (Edson nd Vinson, 1977). In ddition to orl uptke, therefore, direct bsorption of nutrients probbly plys n importnt role in nourishment of C. congregt during lrvl development. The nl vesicle of C. congregt is withdrwn towrd the end of the second stdium nd the mndibles become sclerotized in preprtion for incising the host integument followed by emergence. The reltionships between blood protein nitrogen nd trehlose levels in norml nd prsitized M. sext lrve on the vrious diets re strikingly similr to the nutrient intke rrys demonstrting the reltionships between protein nd crbohydrte consumption (Thompson et l., 25). ltertions of nutrient intke s result of prsitism, therefore, my be n dptive strtegy by C. congregt, where

8 632 S. N. Thompson, R.. Redk nd L.-W. Wng Tble 5. NOV summry for models estimting the reltionship between blood protein nitrogen nd trehlose levels (mg per insect) nd non-emerged prsite burden nd biomss Prsite burden Source d.f. Men squre F vlue Probbility r 2 Model < Error Model prmeter estimtes Stndrdized Stndrd regression error of Vrible coefficient coefficient Trehlose Nitrogen Trehlose Nitrogen Intercept.4779 Prsite biomss Source d.f. Men squre F vlue Probbility r < Model Error Model prmeter estimtes Stndrdized Stndrd regression error of Vrible coefficient coefficient Trehlose Nitrogen Trehlose Nitrogen Intercept resultnt blood metbolite levels re closest to optiml for supporting prsite growth nd development on specific diets. The host blood metbolite rry, the reltionship between blood protein nitrogen nd trehlose levels, my represent rule of compromise for the prsite, somewht nlogous to the rules of compromise pprent from nutrient intke rrys. Rules of compromise define how nimls regulte diet consumption, post ingestive responses nd development time to ccommodte imblnces in dietry nutrient composition (Rubenheimer nd Simpson, 1999; Thompson et l., 25). Here, the rule of compromise involves the mnner in which prsitism ffects host feeding nd physiology to influence blood metbolite levels. lthough the host blood metbolite rry for prsitized lrve reflects nutrient vilbility rther thn nutrient uptke by prsites, it is similr to the nutrient intke rrys typiclly observed during development of specilist feeders. In these cses, insects will suffer lrge shortfll of deficient nutrient to void consuming smll surplus of n excessive nutrient, s the nutrient rtio shifts wy from the most optiml rtio. C congregt is reltively host specific, hving been reported from severl host species, ll restricted to the fmily Sphingide (Gilmore, 1938). Host specificity is, in prt, defined by the bility of C. congregt to modify the feeding nd physiology of its host, M. sext, resulting in the most suitble milieu for supporting successful development of the immture prsitic stge. Other prsitoids my bring bout different effects on the feeding nd physiology of M. sext, or other host species, likewise producing suitble nutritionl environments but reflecting different developmentl strtegies nd host specificity. Prsites developing in M. sext lrve include those tht successfully emerge from hosts to pupte nd complete development nd those tht fil to emerge. Over wide rnge of finl host size there is mximum number nd totl biomss of prsites tht emerge from individul host lrve. In our study, these were pproximtely 1 prsites nd 1 mg totl biomss. This burden of prsites is considerbly lower thn the pproximte 2 mximum number reported by lleyne nd Beckge (1997) for M. sext lrve similrly prsitized by C. congregt but mintined on the whet germ-bsed rering diet. Lrve purportedly grow s well on the chemiclly defined diet s on the rering diet (hmd et l.,

9 Host nutrition medites prsite development ), but specific differences in nutrition my ccount for the different results for emerged prsites. Depending on diet, the number nd totl biomss of non-emerged prsites my exceed those of prsites tht successfully emerge. This phenomenon, lrge number of prsites filing to emerge, hs been described in both field collected (Fulton, 194; Thurston nd Fox, 1972) nd lbortory rered M. sext lrve (Brbos et l., 1991; lleyne nd Beckge, 1997). Hosts rered on three diets,.5c:1.5s, 1.C:1.S nd.5c:1.5s, supported mximl nd similr numbers nd biomss of emerged prsites. Hosts on the 1.C:1.S supported the lrgest prsite numbers nd biomss of prsites, including both those tht emerged nd those tht filed to emerge. The sme diet supports the gretest mss gin by prsitized lrve (Thompson et l., 25), nd hs the rtio of protein nd crbohydrte selected by prsitized lrve when offered choice of high protein nd high crbohydrte diet (Thompson et l., 21). The effects of dietry nutrient rtio on prsite burden nd biomss re supported by dt showing the influence of blood nutrient levels on prsite prmeters. Different responses of emerged nd non-emerged prsites to blood nutrient levels were redily pprent from the models nd contour mps illustrting these reltionships. However, the specific nutrient levels supporting the gretest prsite burden nd biomss reflect protein:crbohydrte rtios vrying between pproximtely 1 nd 1.5, within the rnge of the dietry nutrient rtios supporting the gretest prsite burden nd biomss, whether emerged or non-emerged. Future investigtions will exmine how host protein nd crbohydrte intke re prtitioned into host nd prsite growth. Our studies estimte nutrient concentrtions nd levels t pproximtely hlf wy through the fifth stdium only nd my not be reflective of nutrient vilbility throughout the entire period of prsite development. The concentrtions of vrious blood metbolites in prsitized lrve my chnge over time (Vinson nd Iwntch, 198). Studies by others demonstrte tht in M. sext lrve prsitized erly in the fourth stdium, blood protein concentrtion continuously increses during the fifth stdium, lbeit t lower rte thn in norml lrve, nd reches pproximtely 11 mg ml 1 when feeding stops (Beckge et l., 1989; Beckge nd Knost, 1993). The protein concentrtion then declines to pproximtely 7 mg ml 1 when prsites emerge. In those studies, the protein concentrtion t mid-fifth stdium, pproximtely 9 mg ml 1, is similr to the concentrtion we observed t the sme point of development in prsitized lrve on the.5c:1.5s diet (Thompson et l., 25), lthough the ltter diet contins bout 2% less protein thn the whet germ diet on which lrve were fed in the bove investigtion. Some investigtors hve suggested tht prsite emergence is limited becuse M. sext host lrve become exhusted of blood nd ft body, concluding tht prsites filing to emerge simply hve not consumed sufficient nutrients (Bentz nd Brbos, 199; lleyne nd Beckge, 1997). Some of our results pper to support this conclusion. The observtion, for exmple, tht the burden, biomss nd proportion of nonemerged prsites to totl prsites re highest in the lrgest hosts fed the 1.C:1.S diet, tht lso hve the gretest proportion of totl prsite biomss to finl host mss. Our erlier results demonstrting tht prsitized lrve feeding on this diet significntly increse diet consumption (Thompson et l., 25) would suggest tht lrve ccommodte ny incresed demnd for nutrients. The present finding tht nonemerged prsite burden increses long with incresed protein consumption would lso rgue ginst this. If nutrition were limiting, we would predict tht hosts mintined the 1.C:1.S diet, but with low prsite burden, would hve fewer nd lower proportion of prsites filing to emerge. To test this, we exposed M. sext lrve individully to C. congregt femles nd removed hosts following single oviposition encounter. fter completing development, the numbers of emerged nd non-emerged prsites were determined. Prsite burdens generlly rnged from 5 to 2, much lower thn the burdens for superprsitized hosts mintined on the better experimentl diets. The wide rnge of prsite burden my in prt be due to the mount of time femles spend during single oviposition, which vries from one to severl seconds. Regrdless of the prsite burden, however, in every cse t lest hlf of the prsites filed to emerge. This is consistent with, lthough somewht higher thn, the 25% verge nonemerged prsites reported by Thurston nd Fox (1972) for field collected host lrve with prsite burdens of 5 1. Persusive evidence ginst the hypothesis tht nutrition limits prsite emergence is the observtion tht mny nonemerged prsites re mture second instr lrve nd pper morphologiclly indistinguishble from lrve tht do emerge. The similr mss of emerged nd non-emerged prsites found in the present study strongly suggests they were similrly nourished. In contrst to results of some others (Beckge nd Riddiford, 1982; lleyne nd Beckge, 1997), we did not observe ny significnt decrese in the size of individul prsites with incresed prsite burdens. In our studies, however, differences in prsite burden were chieved by mintining hosts on different diets, while in the studies of lleyne nd Bekge (1997) were found highly vrible prsite burdens nd individul prsite mss in hosts mintined on the sme diet. dditionl study is necessry to ssess the potentil role of nutrient depletion in prsite filure to emerge. Experiments, for exmple, involving dministrtion of supplementl nutrients through injection into hosts fter cesstion of host feeding my prove useful for indicting the potentil of nutrition to lter prsite success. lterntely, prsitized hosts might be mnipulted in mnner tht extends the feeding period nd totl nutrient consumption. t this time, however, we believe tht other fctors re probbly involved in determining prsite emergence, or filure to emerge. Host nutrition my medite prsite development nd success through effects on host endocrinology, which prsitism disrupts. bnormlly elevted blood concentrtion of juvenile hormone nd lower thn norml 2-

10 634 S. N. Thompson, R.. Redk nd L.-W. Wng hydroxyecdysone concentrtion occur during prsitism of mny insects, including M. sext (Beckge nd Gelmn, 21; Cole et l., 22). Elevted juvenile hormone explins the decresed growth nd delyed development of prsitized host lrve nd ultimtely their filure to pupte. Furthermore, juvenile hormone probbly influences prsite development, s ppliction of juvenile hormone or methoprene, juvenile hormone gonist, to the cuticle of intct prsitized lrve prevents prsite emergence (Beckge nd Riddiford, 1982). Host lrve probbly disply vrying ptterns of developmentl hormone concentrtions depending upon nutritionl sttus, s reflected by the effects of nutrition on host size (Nijhout, 1994) s well s prsite burden. In norml lrve, such responses ensure tht moulting nd metmophosis occur t pproprite times under specific nutritionl conditions. Under suboptiml nutritionl conditions, juvenile hormone concentrtions my remin high for longer periods, delying development while lrve grow to n dequte size. M. sext lrve fed only sucrose during the first 3 dys of the fifth stdium disply delyed development nd metmorphosis in response to slower thn norml decrese in juvenile hormone synthesis (de l Grz et l., 1991). In prsitized insects, prticulrly in host lrve on suboptiml diets, ltered hormone concentrtions my desynchronize the development of the prsite. In the cse of M. sext prsitized by C. congregt juvenile hormone concentrtion begins to decrese few dys before prsite emergence. If juvenile hormone is too high t the time prsites re redy to begin emergence nd moulting, prsites my become unresponsive nd their further development permnently delyed. Thus, the distribution of emerged nd non-emerged prsites my in prt reflect differentil response mong prsites to hormone levels in hosts on the vrious diets. significnt loss of prsite resources ppers to chrcterize the nutritionl nd developmentl interctions between C. congregt nd M. sext. First, lrge number of prsite eggs, or lrve, probbly erly instrs tht re no longer pprent upon dissection, fil to develop. We ssume becuse ll host lrve were rered under the sme nutritionl conditions, nd were of the sme developmentl stge nd size t the time of oviposition, tht ll re of comprble nutritionl qulity. Furthermore, becuse ll hosts were prsitized in like mnner, we ssume tht similr numbers of prsite eggs re deposited in most host lrve. lthough the initil number of eggs ws not determined, the difference in totl prsite burden between the best nd poorest diets suggests tht if the bove ssumptions re ccurte lrge portion of eggs fil to develop. lthough ovicide, the destruction during superprsitiztion of one prsitoid s eggs by nother prsitoid, usully through use of the ovipositor, might provide n explntion, ovicide is unknown in Cotesi spp. or other gregrious endoprsites. Second, of the eggs tht develop into mture lrve, lrge portion fil to emerge. ll of this suggests tht C. congregt exhibits high degree of imprecision in determining n optiml number of eggs to deposit in host lrve. This my in prt be becuse C. congregt oviposits eggs quickly, nd my therefore be unble to llocte eggs precisely. lso, ssessing the optiml egg number required to efficiently relize the gretest reproductive potentil or rte of gin of fitness is often compromised by fitness penlties for depositing too few eggs (Godfrey, 1994). Fitness penlties for ovipositing more eggs thn successfully develop relte to the prsite s egg number, or cpcity to produce eggs, nd the scrcity of host individuls to prsitize (Weisser nd Houston, 1993). C. congregt, pro-ovigenic koinobiont (Godfrey, 1994; Quickie, 1997) eclosing to the dult stge with very lrge numbers of eggs, so tht the fitness penlties for ovipositing too mny eggs my be miniml if hosts re scrce. C. congregt prsitizes vriety of Sphingid moth lrve (Gilmore, 1938), which generlly disply wide dispersl ptterns. M. sext deposit eggs in ptches, but ptches cn be widely distributed nd individul grvid femle moths my fly continuously over n entire night (Gilmore, 1938b). In cultivted cotton, moths ly one to five eggs on individul plnts nd femles hve s mny s 2 eggs. In the cse of wild host plnts, principlly solnceous species, ptches would be even more widely distributed. Becuse over rther wide vrition in nutritionl conditions the number nd biomss of emerged prsites is nerly constnt, deposition of excess eggs by C. congregt my not be dptive, but neither my it be detrimentl. The potentil dptiveness of superprsitism my differ depending on whether superprsitism is due to multiple prsitiztion by n individul prsitoid, self-superprsitism, or is due to conspecific superprsitism, prsitiztion by more thn one prsitoid (vn lphen nd Visser, 199). Our csul observtion during the present study is tht superprsitism ws principlly conspecific. In either cse, however, superprsitism my be dptive for n individul prsitoid competing for scrce hosts. References hmd, I. M., Wldbuer, G. P. nd Friedmn, S. (1989). defined rtificil diet for the lrve of Mnduc sext. Entomol. Exp. ppl. 53, lleyne, M. (1995). ltertions in host growth nd metbolism of Mnduc sext lrve prsitized by Cotesi congregt. MS thesis, University of Cliforni, Riverside, C, US. lleyne, M. nd Beckge, N. E. (1997). Prsitism-induced effects on host growth nd metbolic efficiency in tobcco hornworm lrve prsitized by Cotesi congregt. J. Insect Physiol. 43, Bker, F. C., Tsi, L. W., Reuter, C. C. nd Schooley, D.. (1987). In vivo fluctutions of JH, JH cid, nd ecdysteroid titer, nd JH esterse ctivity, during development of fifth stdium Mnduc sext. Insect Biochem. 17, Brbos, P., Gross, P. nd Kemper, J. (1991). Influence of plnt llelochemicls on the tobcco hornworm nd its prsitoid, Cotesi congregt. Ecol. 72, Beckge, N. E. nd Gelmn, D. B. (21). 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