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Animal Behaviour 81 (2011) 993e999

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Animal Behaviour

journal homepage: www.elsevier .com/locate/anbehav

Nutrient regulation in a predator, the wolf spider Pardosa prativaga

Kim Jensen a,b,c,*, David Mayntz c,d,1, Søren Toft c,1, David Raubenheimer e,2, Stephen James Simpson b,3

aDepartment of Zoology, University of Oxfordb School of Biological Sciences, University of SydneycDepartment of Biological Sciences, Ecology and Genetics, University of AarhusdDepartment of Genetics and Biotechnology, University of Aarhuse Institute of Natural Sciences, Massey University

a r t i c l e i n f o

Article history:Received 9 November 2010Initial acceptance 4 January 2011Final acceptance 28 January 2011Available online 8 March 2011MS. number: 10-00778

Keywords:dietgeometric frameworklipid:protein ratioLycosidaenutrient balancingPardosa prativagaperformancewolf spider

* Correspondence and present address: K. JensenExeter, Cornwall Campus, Daphne du Maurier, Penryn

E-mail address: kim.jensen@biology.au.dk (K. Jens1 D. Mayntz and S. Toft are at the Department of Bio

Genetics, University of Aarhus, Building 1540, 8000 Å2 D. Raubenheimer is at the Institute of Natural

Albany, Private Bag 102 904, North Shore Mail Centre3 S. J. Simpson is at the School of Biological Sci

Heydon-Laurence Building A08, NSW 2006, Sydney, A

0003-3472/$38.00 � 2011 The Association for the Studoi:10.1016/j.anbehav.2011.01.035

Nutrient balancing is well known in herbivores and omnivores, but has only recently been demonstratedin predators. To test how a predator might regulate nutrients when the prey varies in nutrientcomposition, we restricted juvenile Pardosa prativaga wolf spiders to diets of one of six fruit fly,Drosophila melanogaster, prey types varying in lipid:protein composition during their second instar. Wecollected all fly remnants to estimate food and nutrient intake over each meal. The spiders adjusted theircapture rate and nutrient extraction in response to prey mass and nutrient composition irrespective ofenergy intake. Intake was initially regulated to a constant lipid plus protein mass, but later spiders fed onprey with high proportions of protein increased consumption relative to spiders fed on other prey types.This pattern indicates that the spiders were prepared to overconsume vast amounts of protein to gainmore lipids and energy. The spiders also regulated protein after ingestion, and ingested protein wasincorporated less efficiently into body tissue when the prey was protein rich. Despite both pre-and postingestive nutrient regulation, the body lipid:protein compositions of the spiders were highlyaffected by the nutrient compositions of their prey, and growth in carapace length and lean body massincreased with increasing prey protein:lipid ratio. Our results demonstrate that prey nutrient compo-sition affects these predators, but also that the spiders possess behavioural and physiological adaptationsthat lead to partial compensation for these effects.� 2011 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved.

In contrast to herbivores (Behmer 2009; Felton et al. 2009) andomnivores (Raubenheimer & Jones 2006; Lee et al. 2008), predatorshave traditionally been thought not to balance nutrient intake. Thisbelief has partly been based on the assumption that animal tissueas a food source varies little and is nutritionally balanced (Westoby1978; Stephens & Krebs 1986; Galef 1996). In addition, predatorsare generally considered energy limited, and the amount of preya predator can catch is typically considered the limiting factor forpredator performance (Kessler 1971; Riechert 1992; Wise 1993).Regarding the first assumption, chemical analysis of invertebrateprey has revealed remarkable variation in nutrient compositionamong species (Elser et al. 2000; Fagan et al. 2002), and evenwithin

, Biosciences, University of, TR10 9EZ, U.K.en).logical Sciences, Ecology andrhus C, Denmark.Sciences, Massey University,, Auckland, New Zealand.ences, University of Sydney,ustralia.

dy of Animal Behaviour. Published

species, nutrient composition may vary considerably depending onfood source and feeding state (Lee et al. 2002; Simpson et al. 2002;Mayntz et al. 2005; Raubenheimer et al. 2007; Salomon et al. 2008).It is probably true that predators are often energy limited, butalthough a predatormay face shortage of prey for extended periods,it would benefit from nutrient balancing during times of plenty.Nutrient balancing would also be of benefit in cases where thequalitative nutritional requirements of the predator change duringdevelopment. In such cases, regulating energy intake alone mightnot match nutrient demands, for example during periods of fastgrowth.

Greenstone (1979) first suggested that predators may selectfood items according to their nutrient contents. This hypothesis haslater been supported experimentally in fish (Oncorhynchus mykiss:Sánchez-Vázquez et al. 1999; Dicentrarchus labrax: Rubio et al.2003), the ground beetle Anchomenus dorsalis (Mayntz et al.2005), mink, Mustela vison (Mayntz et al. 2009) and the ant-eating spider Zodarion rubidum (Pékar et al. 2010). As many pred-ators are sedentary sit-and-wait hunters, however, balancingnutrients through active prey selection may be less pronounced inthese predators compared to predators that actively pursue theirprey. Sit-and-wait predators will be likely to catch whatever prey

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K. Jensen et al. / Animal Behaviour 81 (2011) 993e999994

comes within their range, and prey would be caught and killed foras long as the predator needed nutrients. Nutrient balancing wouldthen be more likely to occur at later stages, that is, during feeding,digestion and assimilation. In accordance with this, the wolf spiderPardosa prativaga, which is an intermediately mobile sit-and-waitpredator, has been shown to regulate nutrient intake by extractingmore dry mass from a prey item if it contained a higher proportionof a nutrient that was deficient in the previous prey (Mayntz et al.2005).

Here we investigated a different situation where juvenileP. prativaga were provided with prey of a constant lipid to proteinratio during an entire instar of development. We then tested howprey nutrient composition would affect the spider’s body compo-sition and performance. By measuring nutrient extraction overeach meal and by comparing nutrient extraction with nutrientgrowth over the instar, we furthermore investigated all possiblebehavioural and physiological mechanisms the spiders could use toregulate nutrients and reduce nutritional imbalance when feedingon nutritionally imbalanced prey.

METHODS

Diets

Drosophila melanogaster fruit flies of different body lipid toprotein (L:P) ratio were produced by rearing the larvae in sixdifferent media (Table 1). Cultures were held in vials, 3.4 cm indiameter, containing 5 g of dry medium plus water and a few dropsof dissolved yeast. Parental flies (40 � 5) were allowed 5 days in thetubes for egg laying. The temperature was held at 24e26 �C. Onlyfemale flies were used, to reduce the variation in prey nutrientcomposition within diets.

Experimental Set-up

Pardosa prativaga wolf spiders carrying eggsacs were collectedin a wet meadow at Brabrand, Denmark, and brought to thelaboratory. Two days after hatchlings emerged they were gentlyremoved from their mother’s back with a paint brush and distrib-uted to individual translucent plastic vials (diameter 2.0 cm, height6 cm), each with a 1 cm, regularly moistened plaster of Parisbottom and a foam rubber stopper. To ensure that the youngspiders were strong enough to catch and kill experimental fruit fliesfrom the start of the experiment, we fed all hatchlings one Sinellacurviseta collembolan daily from a culture reared on yeast until theymoulted to the second instar. Spiderlings were starved over the5 days following the moult to ensure that they would be hungry

Table 1Growth media and body data of the six nutritionally different Drosophila melanogaster fr

Fly type

LP0.89 LP0.64 LP0.40

Growth medium 1:4 Sucrose:Carolina Pure Carolina 1:9 Casein:CarDry mass (mg) 329�7cb 347�7cd 383�10e

Lean dry mass (mg) 222�5a 253�5b 306�8c

Lipids (%) 32�1.1a 27�1.1b 20�0.8c

Crude protein (%) 37�0.8a 42�0.8b 50�1.2c

L:P ratio 0.89�0.05a 0.64�0.03b 0.40�0.02c

No. of samples 17 20 18

The media were based on Carolina Instant Drosophila Medium Formula 4-24 (BurlingtonNeu-Ulm, Germany) or casein (Sigma C-5890, SigmaeAldrich, Steinheim, Germany) at vafemale flies collected from each medium per feeding day, except a few days when down toare mass based. Different letters indicate significant differences (Student’s t test: P < 0.0

and willing to eat from day 1. The experiment was performed at21e25 �C.

Experimental Procedure

On day 1, the spiderlings from nine mother spiders wereweighed to the nearest mg. The spiderlings within and betweenmothers were then distributed as equally as possible among sevengroups, totalling 27e28 spiderlings per group. One of the groupsformed a start sample where spiderlings were killed by freezingat �18 �C. Spiderlings in the remaining six groups each receivedone of the six different fly types only (Table 1). One fresh fly of theallocated nutritional composition was provided daily to eachspider. Flies that were still alive the next day were replaced. Killedflies were recorded. Their remnants were stored individually in thefreezer for chemical analysis the day after they were presented tothe spiders. A reference sample of 25 flies was collected daily fromeach growth medium to ensure reliable estimates of nutrientcontents of provided flies. On a few days with low fly populationson some media, samples down to 15 reference flies were taken.Spiders were killed on the day of moulting into the next instar byfreezing at �18 �C, and the carapace lengths of all spiders weremeasured with an eye piece scale under a microscope.

Nutritional Analyses and Growth

At the end of the experiment, all spiders, food remnants andreference flies were dried in a vacuum oven over 4 days at 60 �C andweighed. Lipids (L) in each sample were extracted in two 24 hwashes of 2 ml of chloroform, and samples were again dried andweighed. Lipid masses were calculated by subtracting sample lean(lipid extracted) dry masses from sample dry masses. Nitrogencontent was analysed in a combustion analyser (Na 2000, CarloErba, Rodano, Italy). Because of technical problems, 16 spidersacross diets were lost during nitrogen analysis. Food and nutrientintake of each meal consumed in the second instar were calculatedby subtracting the dry masses and nutrient contents left in the flyremnants from the average dry masses and nutrient contents of thecorresponding reference flies sampled on the feeding day. Initialcarapace length, dry mass and nutrient contents of the spiderlingswere estimated from their initial wet masses using linear correla-tions from the spiderlings in the initially killed start sample.Growth in linear body size, dry mass and nutrient masses werethen calculated by subtracting estimated values from thevalues measured at the end of the experiment. Crude protein (P)masses and food energy contents were calculated using standardestimates of 6.25 mg protein per mg nitrogen, and 17 or 37 joulesper mg protein or lipid, respectively (AOAC 2006).

uit fly types used in the experiment

LP0.25 LP0.15 LP0.10

olina 1:4 Casein:Carolina 2:3 Casein:Carolina 3:2 Casein:Carolina352�8d 316�7b 287�7a

303�5c 286�7c 264�7b

14�1.0d 10�0.9e 8�0.7e

58�1.8d 68�2.2e 78�2.2f

0.25�0.02d 0.15�0.02e 0.10�0.01e

20 18 17

, NC, U.S.A.), which was used in its pure form or mixed with sucrose (Fluka, 84097,rying ratios. The fly data are mean � SE from 17 to 20 samples each consisting of 2515 female flies were collected from somemedia owing to shortage of flies. All ratios

5).

(a)

0.6

0.8

1

Prop

orti

on o

f sp

ider

s ki

llin

g th

e fl

y

0.2

0.4 LP0.89 LP0.64 LP0.40 LP0.25 LP0.15

Day 1 2 3 4 5 6 7

0

LP0.10

(b)

0.2

0.25

Lip

id +

pro

tein

ext

ract

ed (

mg)

0.1

0.15 LP0.89 LP0.64 LP0.40 LP0.25 LP0.15

Consecutive fly killed1 2 3 4 5

0.05

LP0.10

Figure 1. (a) Proportion of spiders killing their fly from the first feeding day ondifferent diets and (b) nutrients (lipid and protein) extracted from the first five killedflies per spider (mean � SE). Sample sizes in both figures are 27e28 per group for thefirst feeding and drop during the period as spiders that had moulted are not included.

K. Jensen et al. / Animal Behaviour 81 (2011) 993e999 995

Statistical Analyses

Number of flies killed and intakes of dry mass and energy werecompared across diets using analysis of variance (ANOVA) tests.Nutrient compositions remaining in discarded remnants werecompared to available nutrient compositions in the reference fliesusing post hoc Bonferroni-corrected t tests. To analyse data onnutrient regulation in the spiders, we used the geometric frame-work, which is a state-space modelling approach that was designedspecifically to quantify the main and interactive effects of two ormore nutritional components on intake and postingestive alloca-tion (Raubenheimer & Simpson 1993; Simpson et al. 2004).Cumulative intake from the first five killed flies and the total intakeacross the instar were used to plot lipid versus protein intakearrays. The shapes of such arrays reveal how the intake of onenutrient is prioritized relative to the intake of the other. To test fordifferences in lipid plus protein mass extracted over consecutivemeals, we fitted a line with slope �1 through cumulative lipidversus protein intake for each consumed fly and compared theresiduals across diets using ANOVA tests. A slope of �1 would beconsistent with spiders maintaining a constant nutrient massintake rather than a constant energy intake (where the slopewouldbe �0.5 given that lipid is twice as energy dense as protein). Theutilization efficiencies of lipids and protein were tested usinganalysis of covariance (ANCOVA) tests with nutrient growth as thedependent variable and linear or logarithmic nutrient intake as thecovariate. Final spider body L:P ratios were compared using ANOVAafter arcsine transformation (Zar 1999). Growth in body dry mass,body lean dry mass and carapace length were analysed usingANCOVA tests with spider initial wet mass as the covariate(Raubenheimer & Simpson 1992, 1994; Raubenheimer 1995).Duration of the second instar was compared across diets witha Wilcoxon test. Treatment groups were compared individuallyusing Student’s t test. Where parametric measures were used,residuals were normal or nearly normal. All statistical analyseswere performed in JMP 7.0 (SAS Institute, Raleigh, NC, U.S.A.).

A period is included only if the sample size per group was nine or above.

RESULTS

Diets

The female flies produced on the six different media differedmarkedly in dry mass and body composition (Table 1). Flies raisedon sugar-rich media contained more lipids and less proteincompared to flies raised on more protein-rich media. Fly drymasses were highest on the intermediate media and smaller whenflies were reared on media containing high amounts of protein orlipids (Table 1).

Dry Mass and Energy Intake

The daily probability that a spider would kill the provided fly(Fig. 1a) decreased during the instar for all prey types. Also, thenutrient mass extracted from successive killed flies decreased on alldiets (Fig. 1b). The number of flies killed by spiders in the secondinstar differed across dietary treatments (ANOVA: F5,162 ¼ 3.26,P ¼ 0.008), with flies from media producing smaller dry bodymasses being killed in higher numbers than larger flies (Fig. 2a,Table 1). As a result, total dry mass consumption over the entireinstar did not differ significantly across dietary treatments (ANOVA:F5,161 ¼ 0.10, P ¼ 0.99; Fig. 2b). Total energy intakes (ingested lipidplus protein energy) were highest on the most lipid-rich diets andlowest on the intermediary diets, with significant overall differ-ences (ANOVA: F5,162 ¼ 2.89, P ¼ 0.016; Fig. 2c).

Nutrient Extraction

In almost all cases, the nutrient contents of the discarded flyremnants did not differ significantly from the contents of the liveflies (Bonferroni, t tests: P > 0.05; Fig. 3a), which indicates that thespiders extracted nutrients in proportions that were not differentfrom the proportions in the prey (Fig. 3a). After the first meal,though, remnants of the LP0.89 flies contained significantly lowerL:P ratios than the corresponding reference flies (Bonferroni, t test:P ¼ 0.001; Fig. 3b), indicating that the spiders feeding on the mostlipid-rich flies extracted lipids in a higher proportion than ifextracting at random during their first meal. Spiders feeding on theLP0.40 flies apparently extracted a higher proportion of protein thanthe proportion in the flies over the experiment (Bonferroni, t test:P ¼ 0.027; Fig. 3a), but since no clear patternwas found in the otherdiet treatments this might be a type 1 error.

When protein intake was plotted against lipid intake, we founda distinct pattern of macronutrient regulation (Fig. 4). Over the firstmeals, intake points aligned along a slope that was not significantlydifferent from �1 (ANOVA: P > 0.05), indicating that, at this point,the spiders’ mass intake of the two combined nutrients was nearlyconstant or regulated so thatmass-based excesses and deficits wereequal. During the last meals before moulting, though, protein wasprogressively ingested on the twomost protein-rich diets, which bythe fifth meal bent the intake array outwards about a pivot point atL:P ¼ 0.25, and the array from this meal no longer followed a line

(a)FliesRemnants

0.8

0.4

0.6

1

Fly type

0LP0.89 LP0.64 LP0.40 LP0.25 LP0.15 LP0.10

0.2

(b)

L:P

con

ten

t

0.8

0.4

0.6

1

Flies RemnantsFirst killed LP0.89 flies

0

0.2

Figure 3. Nutrient compositions for the reference flies and the discarded remnants (a)for all fly types over the experiment (mean � SE) and (b) for the most lipid-rich flytype (LP0.89) from the first spider meal in the second instar (mean � SE).

0.4

Fly 3Fly 4

Fly 5*Total*

LP0.64

LP0.89

Lip

id i

nta

ke (

mg)

0.2

Fly 1

Fly 2

LP0.10

LP0.15LP0.25

LP0.40

5

5.5

6P = 0.008(a)

a

No.

of

flie

s ki

lled

4

4.5 bcbc

c

bcab

3.50

1.05P = 0.99(b)

0.2 0.4 0.6 10.8

Dry

mas

s in

take

(m

g)

0.95

1

aa a a

aa

P = 0.016(c)22

0.90 0.2 0.4 0.6 10.8

Ener

gy i

nta

ke (

J)

abc

bc bc

ab

a

16

18

20

0 0.2 0.4 0.6 10.8

Fly L:P content

Protein-rich prey Lipid-rich prey

c14

Figure 2. (a) Total number of flies killed (mean � SE), (b) dry mass extracted(mean � SE) and (c) lipid þ protein energy extracted (mean � SE) by P. prativagaspiders during the second instar. The P values are from ANOVA tests across dietarytreatments. Different letters indicate significant differences between dietary treat-ments (Student’s t test: P < 0.05).

K. Jensen et al. / Animal Behaviour 81 (2011) 993e999996

with a slope of �1 (ANOVA: P < 0.01; Fig. 4), which indicatesincreased food intake by spiders feeding on protein-rich flies rela-tive to spiders on the other diets.

Crude protein intake (mg)

0 0.2 0.4 0.6 0.8 1

Figure 4. Cumulative protein and lipid consumption from successive flies killed byP. prativaga spiders during the second instar (mean � SE). As spiders killed unequalnumbers of flies, only consumption from the first five killed flies plus totalconsumption over the instar is included in the figure. *Arrays that are significantlydifferent from a line with slope �1.

Nutrient Utilization

The lipid utilization plot (Fig. 5a) shows a close to linear rela-tionship between lipids consumed and lipids accumulated in spiderbodies (linear regression: R2 ¼ 0.60, F1,161 ¼ 240.71, P < 0.0001). AnANCOVA on lipid growth with lipid consumption as covariaterevealed no significant covariate*diet interaction (ANCOVA:

F5,161 ¼1.17, P > 0.05). Furthermore, the diet treatment factorremained nonsignificant after we removed the interaction termfrom the statistical model (ANCOVA: F5,161 ¼1.99, P > 0.05). Thisindicates that ingested lipids were stored with nearly equal effi-ciency in spiders across dietary treatments, and the relationbetween lipid intake and lipid body growth could be described asLgrowth ¼ 0.00 þ 0.48 � Lintake.

The utilization plot for protein (Fig. 5b) decelerated at higherprotein intakes and was thus described better by a logarithmic

0.2

0.1

(a)

LP0.25

LP0.40

LP0.64

LP0.89

0.1 0.2 0.3 0.4

Lip

id g

row

th (

mg)

0

LP0.10

LP0.15

Lipid intake (mg)

0.2

0.4

(b)

LP0.10 LP0.15 LP0.25 LP0.40

LP0.64

LP0.89

Crude protein intake (mg)

0.2 0.4 0.6 0.8 1

Cru

de

pro

tein

gro

wth

(m

g)

0

Figure 5. Utilization plots of growth versus consumption over the second instar byP. prativaga spiders on the six diets (mean � SE). (a) Lipid growth versus lipidconsumption. (b) Protein growth versus protein consumption.

P < 0.0001(a)

f

0.6

L:P

con

ten

t

0.2

ab

c

d

e0.4

0.4 P < 0.0001(b)

a

0 0.2 0.4 0.6 10.8

Lean

dry

mas

s gr

owth

(m

g)0.2

0.3 ababc

bc

cd

d

0.35 P = 0.037(c)

0 0.2 0.4 0.6 10.8

0.25

0.3

ab

a

ab ab

b

b

Car

apac

e le

ngt

h g

row

th (

mm

)

0.2

Fly L:P content

Protein-rich prey Lipid-rich prey

0 0.2 0.4 0.6 10.8

Figure 6. (a) Body L:P composition (mean � SE) of P. prativaga spiders after the moultto the third instar. (b) Lean body mass growth (mean � SE) and (c) carapace lengthgrowth (mean � SE) of P. prativaga spiders from the start of the second instar to thestart of the third instar. The P values are from (a) ANOVA or (b,c) ANCOVA tests acrossdietary treatments. Different letters indicate significant differences between dietarytreatments (Student’s t test: P < 0.05).

K. Jensen et al. / Animal Behaviour 81 (2011) 993e999 997

model between protein intake and protein growth than by a linearrelationship: Pgrowth ¼ 0.11 þ 0.030 � lnPintake (R2 ¼ 0.60). When anANCOVA on protein growth was performed with protein intake asthe covariate, we found a significant covariate*diet interaction(F5,147 ¼ 3.56, P ¼ 0.005) indicating that protein utilization variedacross diets, with lower protein utilization on themost protein-richdiets (Fig. 5b).

Body Composition and Growth

The ratio of lipid to protein (L:P ratio) in spider bodies at the endof the instar was highly affected by diet (ANOVA: F5,139 ¼ 108.81,P < 0.0001). Spider bodies reflected the fly nutrient composition(Table 1), and the L:P ratio of the spiders increased stepwise aboutfourfold from the most protein-rich fly type to the most lipid-richfly type (Fig. 6a).

Spider survival was very high and in no treatment didmore thanone spider die during the experiment. The duration of the secondinstar lasted 13.1 � 0.11 (mean � SE) days and did not differsignificantly across dietary treatments (Wilcoxon: c5,162

2 ¼ 2.77,P ¼ 0.74). The total drymass growth of spiders likewise did not varysignificantly across dietary treatments (ANCOVA: F5,162 ¼ 0.38,P ¼ 0.86). However, spiders fed on more protein-rich flies hadsignificantly more growth in lean body mass (ANCOVA:F5,162 ¼ 11.20, P < 0.0001; Fig. 6b) and carapace length (ANCOVA:F5,161 ¼ 2.44, P ¼ 0.037; Fig. 6c) compared to spiders fed more lipid-rich flies. The increase in carapace length spanned from 21 � 3%(mean � SE) in spiders fed the most lipid-rich (LP0.89) flies to33� 3% (mean � SE) in spiders fed the protein-rich LP0.15 flies.

Summary of Nutrient Acquisition and Allocation Responses

Figure 7 summarizes the relationship between nutrients avail-able to the spiders (i.e. the nutritional value of flies provided),

nutrients potentially available in killed prey, nutrients actuallyextracted from the killed prey and nutrients allocated to growthduring the instar. Spiders were provided with more flies than theykilled and did not completely consume all nutrients from killedprey. However, the nutrient compositions available in the killedflies generally did not differ from the nutrients actually extracted.

DISCUSSION

Our study showed clear effects on body composition and growthin juvenile P. prativaga during one instar of development after we

0.8LP0.89

LP0.64

LP0.40

Flies provided

Flies killed

Lip

ids

(mg)

0.4 LP0.25

LP0.15

Extracted

Grown

Crude protein (mg)

0.4 0.8 1.2 1.60

LP0.10

Figure 7. Arrays summarizing nutrient acquisition and allocation by P. prativagaspiders over the second instar. All values are mean � SE. The outermost array showsthe lipid and protein masses available in the provided flies. The next array shows thenutrient masses available in the flies that were killed by the spiders. The third arrayshows the nutrient masses that were extracted from the flies. Finally, the innermostarray shows lipid and protein growth by the spiders. Reference fly means from theactual feeding days are used in all calculations.

K. Jensen et al. / Animal Behaviour 81 (2011) 993e999998

added pure protein (casein) or sugar to the diet of their prey (Fig. 6).Earlier studies have shown that enriching the growth medium of D.melanogaster prey with dog food also increases performance oftheir wolf spider predators (Mayntz & Toft 2001; Wilder & Rypstra2008). High rates of growth are important performance estimatesfor several reasons, including escaping similar-sized predators(Borre et al. 2006; Mayntz & Toft 2006), being able to reproduceearlier or having higher reproductive potential (Petersen 1950;Kessler 1971; Vollrath 1987; Honek 1993) and being favoured incourtship (Maklakov et al. 2004; Lomborg & Toft 2009). Theincreased growth on low L:P ratios indicates that wolf spiders,like many other predators, are adapted to a diet rich in protein(White 1978). However, our experiment also showed evidence thatthe spiders possess both behavioural and physiological mecha-nisms for nutrient regulation, which partially compensate for thecosts of feeding on nutritionally imbalanced prey.

Probability of Killing Prey

Optimal foraging theory assumes that the feeding behaviour ofpredators has evolved under a strong selection pressure to maxi-mize capture rates and energy intake (Stephens & Krebs 1986), andaccording to this it would be plausible to predict that predatorswould have evolved to kill all prey they could catch. This was notthe case. Spiders did not always kill flies and did not exhibitsuperfluous killing as has sometimes been reported for otherpredators (Maupin & Riechert 2001; Fantinou et al. 2008). Thespiders killed at a high rate in the early days of the experiment(Fig. 1a) when their need for nutrients was highest. Later, decliningnutrient masses were extracted from the flies (Fig. 1b), and preywas more often left alive (Fig. 1a), even though the experimentalsetting allowed easy capture. The different number of prey itemskilled over the instar depending on fly type (Fig. 2a) suggests thatthe spiders varied the number of prey they killed depending ontheir need for food mass (Fig. 2b) and nutrients (Fig. 4).

Total Intake and Nutrient Intake Arrays

As a result of differences in the number of flies killed, the totalmass of macronutrients consumed did not differ across dietarytreatments, whereas total energy gain varied widely (Fig. 2c).Hence, we cannot reject the simple hypothesis that regulation ofdrymass intake alone determined food intake. The intake arrays forprotein and lipid at the beginning of the instar reflect this

possibility. The concave array that developed towards the end ofthe instar, however, indicates that nutrient-specific intake regula-tion became apparent at this point. Probably, the different dietnutrient compositions began to have pronounced physiologicaleffects at this point. Increasing intake on nutritionally extremediets resulted in spiders gainingmore of the limiting nutrient in thediet, in this case lipids, at the cost of ingesting a greater amount ofthe nutrient in excess, in this case protein. The ground beetleA. dorsalis showed a similar intake pattern when restricted tonutritionally fixed foods after winter hibernation. They also formedan initial intake array with a slope of �1 and later developed anoutwards curvilinear intake array, as beetles providedwith protein-rich food increased intake relative to beetles on the other diets(Raubenheimer et al. 2007). The starvation period before feeding inour experiment and the hibernation period might explain thesimilar results. The pivot point about which the nutrient intakearray opens out over time (Figs 4, 7) coincides with the LP0.25 diet.This diet also corresponded to the point of deflection in the proteinutilization plot (Fig. 5b), beyond which excess ingested protein wasconverted to tissue growth at a lower rate. These two findingspossibly reflect that the LP0.25 diet contains a nutrient compositionthat balances the two nutrients, so both are most efficiently utilizedby the spiders (Simpson & Raubenheimer 1995).

Nutrient Extraction

During the first meal, the spiders provided with the most lipid-rich flies (LP0.89) extracted a higher proportion of lipids than theproportion in the flies (Fig. 3b). Lipids in these flies were veryabundant which might have enabled selective lipid extraction, andthe spiders possibly extracted lipids selectively to restore energystores that were depleted during the previous moult and days ofstarvation. This pattern corresponds to the pattern found in A.dorsalis after winter hibernation: beetles given the chance tochoose their diet first selected lipid-rich food to fill up lipid storesbefore foraging selectively for protein (Raubenheimer et al. 2007).Whereas the sessile spider Stegodyphus lineatus was shown toredress nutritional imbalance by extracting nutrients selectivelyfrom the prey (Mayntz et al. 2005), P. prativaga was instead shownto consume higher masses from nutritionally complementary preythan from nutritionally similar prey (Mayntz et al. 2005). This is inaccordancewith our general finding that P. prativaga does not showextensive selective nutrient extraction but rather regulates massintake to balance the combined intake of lipids and protein.

Postingestive Nutrient Regulation

Our results show that lipids were incorporated into body tissueswith nearly similar efficiency regardless of prey nutrient composi-tion (Fig. 5a), and body lipid growth was therefore closely linked tolipid intake. This is in accordancewith thefinding thatmetabolism isnot raised when these spiders feed on lipid-rich prey (Jensen et al.2010). Protein utilization, however, varied across diets with lowerutilization efficiency above a certain protein intake (Fig. 5b).Postingestive regulation of protein has been shown before in cats,Felis catus (Russell et al. 2002, 2003) and in some herbivores(Simpson et al. 2002; Lee et al. 2004, 2006) and probably reflectsa pronounced ability to use amino acids as a source of metabolicenergywhile voiding the aminogroups (Zanotto et al.1993). Perhapsthe ability to overingest protein as a source of energy is common inpredators as an adaptation to feeding onprotein-rich foods. This is incontrast to herbivores and omnivores, which are not willing tooveringest protein (Raubenheimer & Simpson 1997; Raubenheimer& Jones 2006). As a consequence of the lower rate of protein utili-zation for growth on high-protein diets, the growth array

K. Jensen et al. / Animal Behaviour 81 (2011) 993e999 999

approaches a vertical slopewithprotein growthbeingmore stronglyconserved across diets than lipid growth (Fig. 7). The spiders did notshow postingestive regulation where nutrient growth pointsconverge around a common growth target as seen in herbivores andomnivores (Simpson & Raubenheimer 2000, 2001). As spiders areadapted to periods of starvation (Wise 1993), they are likely to haveadapted to utilizing the nutrients they ingest and waste as little aspossible to avoid the risk of lacking these nutrients in the future.

Conclusions

In summary, our experiment showed that P. prativaga wolfspiders regulated nutrients at several levels of food handling.Despite this regulation, however, spider body composition washighly affected by the nutrient composition of the prey. Thisincomplete regulation may be explained as an opportunisticfeeding strategy that has evolved as an adaptation to unpredictableprey availability. Spiders may thus have adapted to acquiring andmaintaining nutrients when these are available in order to surviveand thrive in periods of prey shortage.

Acknowledgments

This study was supported by a grant from the UK Biotechnologyand Biological Sciences Research Council. D.M. was in receipt ofa grant from the Danish Research Council. D.R. is part-funded bythe National Research Centre for Growth and Development,New Zealand. S.J.S. was in receipt of a Federation Fellowship andcurrently a Laureate Fellowship from the Australian ResearchCouncil.

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