Posted on Authorea 21 Jan 2020 | CC BY 4.0 | https://doi.org/10.22541/au.157963861.14279623 | This a preprint and has not been peer reviewed. Data may be preliminary. o,te,d urtoa hlegssaefo raimt ueogns?D raimadsuperorga- and and individual superorganism. organism If trade-offs? the Do nutrition-mediated in superorganism? same individuals labour the to face the amongst organism and/or – divided from optima compounded etc.) is nutritional scale are cells, different organisms challenges have but cooperating ‘unitary’ nutritional nism of superorganisms, in 2009). do top by cells Wilson, on then, faced (H¨olldobler organisms amongst How, & also division) (cooperating divided are complexity line is organism of germ that or level labour, – cell of additional (somatic division any the reproductive reproduce by of synergistic the not challenges by a characterized do in nutritional are operating The individuals Superorganisms species some same 1989). that the Sober, of such & individuals Wilson be of S. to groups (D. tend are way insects, and eusocial understood, proteins, like well Superorganisms, as are 1999). trans- such storage, Raff, that quantity, or & mechanisms tissue, high (Conlon molecular new conserved in The maintenance, highly or- ingested 2009). into and Mayntz, macronutrients macronutrients maintenance all the & incoming for by are Simpson, in late necessary faced used (Raubenheimer, variation are fats agents, nutrients and with and many selective consumed dealing carbohydrates, While and majority thus 2001). challenges, the Harshman, and consequential & growth, processes, most Zera the organismal 1992; (Stearns, among most ganisms is of nutrients organizer of upstream availability an is Nutrition INTRODUCTION of preference only and the complex between are connection demographics the and 3) success and associated colony may are diet, affect this that combinations balanced understood. and trade-offs nutrient a be nutrients, nutrient how to are for same and beginning there the selection nutrients 2) by to different levels, controlled for leading colony organizational are foragers life’s maximized likely size across that trade-offs, superorganism size diet life-history for and the with basis organism with molecular 1) maximized aligned common were totally these that: a not of reflect highlight each was results and preference how growth, Individuals Our of forager and function survival Interestingly, quality. a growth. between as diet balance regimes. and a nutrient with diet, was several the different growth corresponded measured in colony with protein preference and Optimal of months amounts forager available. increasing was 8 whether with food over size much examined for in and increased development (superorganisms) colony caste, colonies measured worker and (workers) We the growth. of into traits frequency) phenotypic and ( species composition ant in common (both two used we study, this In Abstract 2020 5, May 3 2 1 Scherber Guti´errezYeisson colony investment ant and intake superorganism: macronutrient the of survival and Growth Westf¨alische Wilhelms-Universit¨at M¨unster College Earlham M¨unster of University 3 n hi Smith Chris and , 1 ugPhung Tung , 2 2 aadMumma Harald , aisniger Lasius and 1 .neoniger L. 2 re Ambrose-Winters Arden , oasyhwte rnlt aito ntediet the in variation translate they how assay to ) 2 Christoph , Posted on Authorea 21 Jan 2020 | CC BY 4.0 | https://doi.org/10.22541/au.157963861.14279623 | This a preprint and has not been peer reviewed. Data may be preliminary. )cmol emda h lc adnat iepedi uoeadnrhr mrc Koz 2008); (Klotz, America northern and Europe in widespread ant, garden black the species: ant as two termed for simultaneously commonly out 1) carried was procedure experimental The species Study METHODS AND MATERIALS factors. of other colonies optimum among that variation, include worker environmental hypotheses alternative adult current or growth. Other and experience over the physiology insects. own on solitary maintenance their to of based colony on condition respond based tuned aligned favouring ancestral preferences the by more and not from advantageous food build is is the selectively state of preference instead collectors be worker the but may are that workers prefer this Furthermore, conditions), would is – colonies lab preference hypothesis carbohydrates that (under whether alternative hypothesized high We growth test an lab. However, to colony the growth. ratios with in macronutrient lab growth for maximized maximized that preference what ratios colony nutrient (field) the matched natural larvae examined to we transferred workers. isotope Lastly, completely stable in and a longer rapidly using persist worker, more we and would be maintenance, larva would carbohydrates individual, that and protein while of those that growth types hypothesized and between two We (larvae) these trade-offs experiment. individuals through tracer possible growing nutrients of between explore flow is further the colonies To examined insect (workers). colony social in the inherent adult maintain 1). labour of Fig. of crop trade-off tissue, division standing life-history (current mediated A the protein-limited, maintenance nutritionally of and be a demands being tissue) indeed metabolic there (new may the on growth different to premised size between by is due worker limited hypothesis limited example, alternative carbohydrate are For This more development workers. differ). be colony observa- optima may and sociometric that colonies colony worker with – but and consistent diet that is the worker is in hypothesis (i.e., hypothesis hypothesized protein This nutrients alternative We of protein-limited. availability colonies. important both increasing to An are with mani- available size tions. increase the carbohydrate and and both and for number superorganism would protein worker allowed size model is, of colony that a amounts and as 1995) size and ants worker Raubenheimer, ratio understanding used that & the the we in both (Simpson study, issue of present framework unresolved pulation the geometry major In nutritional a development. a is and 2015). employed size growth al., superorganism insect et and social Wills survival individual of 2012; worker of 2009, Fur- adult basis Simpson, differences. nutritional documenting & The nutrient dynamics, (e.g., Dussutour colony other (e.g., foods short-term growth many natural longer-term on Suarez, with used than focus & content or rather studies Smith protein/nitrogen observational previous 2008; largely confound many Suarez, been that thermore, & have Passera, prey) Gadau, workers 1994; studies insect Tillberg, Nalepa, larger previous Anderson, & increased However, of Hunt Smith, 1942). production 1937; 2012; Smith, Goetsch, al., the Smith, 2010; 2002; et & with Heithaus, Wills Hunt, associated 2007; & Schmidt, production Smith, (Bono been 1974; to 1974; castes worker also availability Passera, sexual increase 2018; has resource and does sociometry Abouheif, content increasing (majors/soldiers) availability & of protein/nitrogen to Wheeler, nutrient due Increased studies 2006; Metzl, increased Tschinkel, part 1937; 2015). in & - (Goetsch, in (Smith pattern size least colonies colony worker general at larger the and is a in in workers relationship As individuals of distributions), This others. developing number size 1998). the among (and 1993, with size castes, applied 1987, increases individual Tschinkel, term different also size size, general worker and of colony sociogenesis, a environment as ratios and is the such and ‘Sociometry’ superorganisms, to numbers 1985). with responsive the Wilson, associated is O. metrics that (E. at the development those ‘sociogenesis’ to coordinated limit termed a may specific, level undergo lineage/species individual colonies the insect at Social occurring processes then level. aligned, superorganismal the not are optima superorganism 2 aisniger Lasius L,15)(Fig. 1758) (L., Posted on Authorea 21 Jan 2020 — CC BY 4.0 — https://doi.org/10.22541/au.157963861.14279623 — This a preprint and has not been peer reviewed. Data may be preliminary. otoswr 125 were Portions The hereafter for (H). details), carbohydrate for change 96 high 3 linear was and a Fig. niger (MH), colonies representing (see medium-high levels of doubling (ML), four rough number medium-low had of 125 (L), starting increments P:C carbohydrate a with The low 1:16 levels: as (2). carbohydrate to to three week up to referred had per 1:2 (protein approximately twice frequency from or composition Feeding P:C (1), diet in week 3). every and (Fig. (0.5), frequency one week henceforth) range. (approximately feeding other P:C P:C census were first the as treatments the and abbreviated after The provision treatments ratios, to to growth). assigned diet randomly of of were quantity month study the present documented of the also in choice experiment diet Colonies The our a workers. informed of fed data number those pilot greatest than workers the These ~125 fewer produced of survival. produced 1:3 of provision 1:1 of parts weekly diet of increasing carbohydrate a a ratio with that to fed P:C linearly those a protein near and with increased the 1:2, diet workers) in of a of varying fed number colonies treatments (the where growth three carbohydrates, Colony among with diet. distributed their 2015/16 of were content in colonies months; conducted six was over experiment pilot (Hadley A ggplot2 using data raw Growth Figures the Colony 2013). from (Hope, 2014). calculated Rmisc Brewer, intervals summarized & and were confidence (Neuwirth is 2018) Data RColorBrewer and term order-dependent. Wickham, with each means not (H using 2016) i.e. are the lme) Wickham, dplyr marginal, tests glm, after 2010), and are lm, made Wickham, model tests (e.g. the these were (Hadley models in 2018); all plyr terms Weisberg, in libraries other based & terms all (Fox tables using of variance of library significance of presence “car” the analysis in the assess Type-II tested 2019). to in them. Team, used function Core of were ’Anova’ to (R both tests the not 3.6.2 for square version was simultaneously chi R research treatments 1-d.f. in this the on done of of was aim analysis effects the data the as All study separately to species but each for species conducted the were compare modelling and analysis All Analysis Data was General period overwintering species. post different, ecologically and multiple, prior with days in out used 15 temperature carried of higher experiments The prevent phase transition. temperature acclimatization to gradual An for (polytetrafluoroethylene) allowed 2018. fluon February and with January covered 25 cotton- 10 first were at containing the incubators at walls boxes of in overwintered plastic Box housed emergence ml were water. cotton. the 500 Colonies until with in with provided escape. plugged housed tubes was and were water food colonies test with present, No stoppered were half-filled week. workers tubes first Once test the cohort. mm after workers’ 18x150 egg-laying in initiated housed females were Most Queens 2017. 12, O. July E. 2008; (Klotz, insects 2017. dead and and niger living gardens L. and and lawns honeydew urban/suburban hemipteran in are abundant 1955). diets highly Wilson, are Natural species diets. Both similar 1955). have Wilson, O. (E. America and de qal otedes o eal ntede osiuinadrcpsseTbeS.Fo etvr were leftovers Food S1. Table see recipes and constitution diet the on vitamins also details Insect For were 2008). Germany) diets. Simpson, Dephyte, the & (Methylparaben, to preservative (Dussutour equally solution a added agar and in Germany) sucrose Biomedicals, and MP (Vandersant, powder egg protein, whey casein, aisneoniger Lasius .neoniger L. 78clne e combination). per colonies (7-8 Fg )qen eecletdi ¨ntr(R,Gray uigasnl uta ih nJl 5, July on flight nuptial single a during M¨unster Germany) (NRW, in collected were queens 2) (Fig. ueswr olce nRcmn I,USA uigtonpilflgt nJn 0and 30 June on flights nuptial two during U.S.A) (IN, Richmond in collected were queens μ ° ( C (127.33 l mr,19 nw stetrgasat itiue nnrhr n iwsenNorth Midwestern and northern in distributed ant, turfgrass the as known 1893 Emery, .niger L. ± μ .8m,mean mg, 0.38 n 15 and ) fatfiilde seblw a ucett anancln rwhand growth colony maintain to sufficient was below) (see diet artificial of l .neoniger L. ° ( C .neoniger L. ± ° for C D fioaoi hmclydfie it rprdusing prepared diets defined chemically isocaloric of SD) 8clne e ramn obnto) n 6for 86 and combination), treatment per colonies (8 3 .neoniger L. .niger L. o w n n ots epciey between respectively, months, one and two for ) n o30 to and wnyfu ooiswr monitored were Colonies Twenty-four . ° .neoniger L. for C .neoniger L. a u oconcurrent to due was μ ooiswere Colonies . lqo every aliquot l L. Posted on Authorea 21 Jan 2020 — CC BY 4.0 — https://doi.org/10.22541/au.157963861.14279623 — This a preprint and has not been peer reviewed. Data may be preliminary. oie ota stpclylble aeil ol eesl nertd h ai ehd fSi tal. was et experiment Shik previous of the methods in basic used The integrated. diet unenriched easily ML the be fed The could were levels. materials three isotopic remaining labelled background the isotopically heavy for and Five that the diet. diet, control so control receiving enriched to modified identical) those the otherwise order with groups, (but isotopes in provided treatment unenriched the were two diet an colonies of of receiving eight flow those either of the or the to census of S1) and assigned (Table final once diet randomly the fed enriched were After were isotope treatment time. carbohydrates) ML over for the assayed C in was from castes differentiate different to to order in mass atomic nodrt xmn h o fntinsit ooy eue tbeiooeple it enriched Diets pulse. isotope stable a used we colony, a into nutrients of isotopes stable flow heavy the the lipid examine with and to mass order lean In mass, dry Experiment analyse Pulse distribution. to Isotopes Gamma number with used the Stable function) were by portion (’glm’ divided the function) models sample linear (i.e., (’lm’ the generalized lipids of models using was analysed mass Linear that was the mass width sample. as head dry normalized the content; of were in mass proportion ants lean the and of and Samples mass mass, dry (2009). lean Both Smith mass, extracted). and dry Tschinkel measures, of three gave protocol the used masse we for ether; dried Rosset, with first (Schwander, were samples size the measured ants washing body of maximal (SZX7, (8 number repeatedly the availability of The stereomicroscope to at USA). Technology, due measured a proxy varied (iMT and colony software using a Canada) iSolutions-Lite per workers magnification Q-Imagine, using as eyes young 56X 2000R, the (Retiga at of used across camera taken distance attached width was were an Head images with 2) USA) width 1993). Fig. Olympus, Head These Tschinkel, colony- colouration, 2005). content. 2009; with Chapuisat, lighter lipids Tschinkel, change & and their and & mass nutrition by (Smith to lean health (recognized responsive mass, typically and dry are development width, aggregate, level head in and measured: individually were characteristics, traits worker different Several variable. second and response last the the Phenotype as between treatments) workers Worker the of numbers experienced in had difference that the frequency census the and first with effects variance the fixed of analysis (i.e. as two-way census P:C a by used diet we grouped and growth overall feeding (corAR1) on of treatments structure of effects autocorrelation the assess first-order To a effects for fixed as colonies the (96 added were (the factor was data) random number colonies. number the a census of census workers as and inspection Additionally, of expressed P:C visual was number ). diet initial ID the DebRoy, frequency, after colony as Feeding Bates, identified and approximated died. polynomial, (Pinheiro, (factorial), was third-order that nlme Growth a colonies library dynamics. as excluding growth from latter census, assess ’lme’ monthly to function each used in species, was each 2014) for Sarkar, using & model treatments mixed-effects (FDR). comparisons to linear rate related post-hoc discovery A for as false survival for used colony corrected was treatments? were in function with p-values vary ’G.multicomp’ differences - growth The assess overall (Herv´e, RVAideMemoire 2014). to Did library used 3) treatments? the were and on treatments? ’G.test’) depending with vary (function vary survival G-tests colony growth Did of died. 1) dynamics queen questions: the the colony related Did if Every three (SZX7, 2) only answer stereomicroscope basis. to ‘dead’ a monthly analysed under considered were a were tubes Data test on Colonies the the workers magnification. of inspecting 40X and development visually at The larvae by way weeks. USA) counting non-invasive three Olympus, a by every in months prepared done were 10 was census food over of monitored batches was new colonies and days three after removed o ahcln.Tenme fat e ooyws8.8 was colony per ants of number The colony. each for , > 8 t60 at 48h 15 ° .Det h o asof mass low the to Due C. and N ± 13 .,mean 2.6, we brvaigcro ihCw s h uesrp for superscript the use we C with carbon abbreviating (when C ± D.Lpdetatoswr oeuiggaier after gravimetry using done were extractions Lipid SD). 4 Lasius okr,w xrce apeo ants, of sample a extracted we workers, ± .neoniger L. . (mean 1.8 .neoniger L. ± usto ih colonies eight of subset a , D.Telpdextraction lipid The SD). n 6for 86 and .niger L. en Posted on Authorea 21 Jan 2020 — CC BY 4.0 — https://doi.org/10.22541/au.157963861.14279623 — This a preprint and has not been peer reviewed. Data may be preliminary. il raswr odce ewe a n ue21 ngas an o ohntrlyocrigcolonies occurring naturally both for lawns grassy in 2018 June and May of between conducted conditions for were trials lab 2018)) Field and (Ogle, library field FSA under preference the (worker in Diet caste developmental function using (’dunnTest’ and analysed method. h) were test Benjamini-Hochberg data 96 Dunn the these vs. a variances, using (24 of by differences heterogeneity Time pairwise followed to variable. test Due the response Kruskal-Wallis predictors. in the categorical a caste the as and were excess more larva) time 9% cent vs. sampling had isotope per same overall, heavy atomic the diets, the for the enriched of Ltd., quantity fed cent same colonies (Sercon per in the spectrometer atomic individuals by the mass 15 The divided as ratio colonies. 60 ANCA- isotope) calculated isotope unlabelled Europa was lighter at PDZ enrichment 20-20 the oven a cent Europa to using drying per Facility (relative PDZ atomic analysis Isotope a a following The Stable elemental in h to UK). Davis and 96 at Cheshire, interfaced stored capsules after California analyser aluminium then of again elemental sterile University and sampled in the GSL then h, at homogenized were out were 24 Colonies carried samples (2009)). was for Dried Tillberg frozen procedure. & The same withdrawn, offered Smith the changed. were of was Colonies not methods larvae diet. the was the five (following the and diet and in diet, the the sources workers in carbohydrate five of sources the protein ratio of the P:C of 4% 125 mass) approximately the dry was (by that glucose 0.9% supplemental so to amounted added nitrate ammonium were supplemental USA) quantity Sigma-Aldrich, small a SLBJ0583V, and USA) Sigma-Aldrich, SHBJ1050, of Nitrate Ammonium unlabelled: EB0056; of nitrate-15N, quantity small A followed. were (2018) onsa ahcln.Temdlicue h bi”tp ie,tede :)a xdfco,adcolony and factor, fixed a as P:C) for “Time diet colonies repeated as homogenize the of (18 specified (i.e., to was non-independence type factor order (corAR1) the ”bait” random structure account the in a into included on transformed, take model as either square-root to The ID were colony. model were field they each mixed-effect to colonies at (i.e. a diets counts field P using of constant from analysed sets or and three collected C 3). variances, presented ’ML’ data constant then Fig. preferred either We preference in most had C. line Diet the diets levels or black be separately, horizontal paired P to the all then either diets or - doubled and these vertical above) and constant), for the halved (as C midpoint then colonies (holding the and lab varied used 3) choiceand We were diet constant). P Fig. additional P (see of an (holding diet Levels varied performed were content. we C C established, of and/or been content had P levels probability by P:C equal ant species four an the other the had with at If test for portions points preference food time minutes. all subsequent food 40 all that After and assumed during We point minutes time analysis. that five the then every in discovery. location, placement, included counted of any After not at was were once. portion portion only location food used that food any was at each each seen and at were apart m species ants 1.5 of least at number were the entrances nest Baited entrance. nest (39 Indiana Richmond, 125 in with College Earlham of campus aawr o ttsial nlsddet o apesz.Bt ooissoe h aepeeec to preference same the showed colonies Both Laboratory size. period. time sample every low for counted to were taken due bait were each Pictures analysed at ants time-lapse. statistically all for containers not and application colony hours were Lapse-It 3.5 to data least the attached at and arenas for iPhones foraging minutes using in 5 every filmed placed was were foraging portions then Food laboratory-reared and workers). two hundred on one conducted data. over also with analyse were to trials used preference was Food 2014) al., et (Pinheiro library nlme 13 oprdt niiul nclne e naelddes aheeetwsaaye eaaeyusing separately analysed was element Each diets. unlabelled fed colonies in individuals to compared N .niger L. aeldo naeldguoe(aeld -lcs--3,MB02V naeld D-(+)-Glucose, unlabelled: MBBC0227V; D-Glucose-1-13C, (labelled: glucose unlabelled or labelled C μ ftede o 4h fe hspro,tefo a eoe n admsml fapproximately of sample random a and removed was food the period, this After h. 24 for diet the of L μ otoso l orPCrto a ecie bv)i admodr a lcdna nactive an near placed was order, random in above) described (as ratios P:C four all of portions l tteBtnclGre fM of Garden Botanical the at .neoniger L. ny h itwsmdfidt seti hte : rfrnewsdriven was preference P:C whether ascertain to modified was diet The only. 15 .neoniger L. aeldo naeldamnu irt lble:Ammonium (labelled: nitrate ammonium unlabelled or labelled N ntr(51 unster ¨ | ooyI attp” saoe h le ucinfo the from function ’lme’ the above, As type”. bait / ID colony 5 ° n 3for 23 and ° 75.” 7 57’55.9”N 91.” 84 49’15.2”N .niger L. ° 62.”) n for and 36’22.9”E), ° .neoniger L. 44.”) . mmcocp slide microscope cm 7.5 A 54’47.0”W). .Afis-re autocorrelation first-order A ). oois(..toyasold years two (c.a. colonies .neoniger L. 13 n 1 more 71% and C nthe on ° C Posted on Authorea 21 Jan 2020 — CC BY 4.0 — https://doi.org/10.22541/au.157963861.14279623 — This a preprint and has not been peer reviewed. Data may be preliminary. r rtigse ywres ihwre tmcprcn of of cent enrichment per atomic atomic the worker double with nearly workers, had by ingested first are hr a ttsial infiateeto at ysml iefraoi niheto both of enrichment atomic for time sample by caste of test, effect (K-W significant statistically a was There individuals. Experiment fatter and (Fig. Pulse larger species Isotope to both led Stable for availability) assessed food variables greater dependent (i.e., all frequency, feedings across lower frequent consistent the was more than effect - the different 6) of marginally on direction were effect the significant treatments mass 0.08), statistically dry frequency a and had higher lean only (both for frequency obtained feeding noted results while workers). as addition, the larger But In with to consistent 6). correspond is diets means (Fig. proteinaceous in mass more difference Although dry (i.e., directional and content. the lipid lean P:C, worker width, for diet opposite by head the particular increased was this effect diet the of in above, traits content measured protein the increasing all Overall, for significant were frequency interactions feeding no and and species. frequency, P:C feeding diet only. and by P:C frequency affected feeding significantly by were affected traits results neoniger full measured and L. All species both for S4. in 6 Table separately Fig. in in summarized given are phenotypes are worker in treatments among Differences Phenotypes Worker winter from ( recovery levels and treatment overwintering only two of in with other Colonies differences significant, the to highly to comparisons). was owing relative growth fed post-hoc were reduced both colonies suffering the week in that produced other frequency (H) every The diet fed carbohydrate comparisons. colonies (L, high post-hoc The carbohydrate in low species, S3). diets both Table the 0.005) for 4b, from significant differing statistically (Fig. were growth, interactions frequency lowest significant feeding no and were P:C there diet growth, but overall non-recovery of the analysis from the apparent In were more becoming growth differences on to and effects due likely of Time*Diet frequency species-specific were dynamics of which interactions Additionally, feeding the of the some that time. P:C, time, suggest were over with (diet interactions pronounced changed as significant effect frequency two-way significant, feeding latter fixed and statistically The P:C each was diet S2). species, ( Table model treatments 5, both mixed-effects other For (Fig. all Time*Frequency the to in correction). in relative time) FDR treatment survival decreased and after diet showed by comparisons (L) overwintered only all carbohydrates spent in affected in (months low was 2018 diet April survival 3, the in Colony Only = last the df analysis). and 9.19, the 2017 = in August in included was not growth colony were of census first The Growth and Survival Colony and P variable the to RESULTS recruitment high with responded pair the of colony S1). one (Fig. only diets but C ratios, diet four the .neoniger L. χ okrla asadlpdcnetwr ffce yde : ny hra edwdhwas width head whereas only, P:C diet by affected were content lipid and mass lean worker , 2 .niger L. > ooisps-itr seilydet ieeta uvvl(above). survival differential to due especially post-winter, colonies P 2 ..=3, = d.f. 12, .3 i.4) u ete pce hwda ffc ffeigfeunyo survival. on frequency feeding of effect an showed species neither but 4a), Fig. 0.03, = u nyha it xiie infiatde : rqec neato.In interaction. Frequency x P:C diet significant a exhibited width head only but , P < .0 o both for 0.005 .neoniger L. 13 .niger L. oprdt l te ramnsadhdfiefl higher five-fold had and treatments other all to compared C okrdyms a h nytataetdb ohdiet both by affected trait only the was mass dry worker 15 6 and N rwlre hnclne of colonies than larger grew P < 13 .5 n eimhg abhdae(MH, carbohydrate medium-high and 0.05) 15 ) tcnb oie hticmn nutrients incoming that noticed be can It C). and N .neoniger L. 13 ekn t2h h 4 workers 24h The 24h. at peaking C .neoniger L. edwdhwsntaffected not was width head .neoniger L. P .6and 0.06 = okrdymass dry worker .neoniger L. 15 seemingly , P and N P < < 0.001 P 0.05 13 P (G = < C Posted on Authorea 21 Jan 2020 — CC BY 4.0 — https://doi.org/10.22541/au.157963861.14279623 — This a preprint and has not been peer reviewed. Data may be preliminary. s uvvl n niiultat sz s ii otn)itreti u aa(i.9.Teae of and survival area colony The both intermediate for the P:C 9). of optimum level the the (Fig. detected at have data is For may our intersection we effect. (growth Interestingly, the fitness in colony study. species, equal intersect this both has both content) traits in for for the lipid used traits, lines of P:C colony-level each The and vs. that particular individual assuming (size a intersect. investment, both for values traits optimal when that namely, the P:C individual size such optima, represents differing different individual and 1, intersection have over for survival) traits Fig. values as different in when well trait depicted vs. exist as representing Trade-offs is P:C. survival, trade-offs lines optimal nutrient and the single of growth a schematic colony is visual there for of A trait trade-offs size storage. nutrient the colonies. lipid of is and within and evidence and determination size growth show caste colony colony also and constrains that We size likely Europe hypothesis of protein central our regulator of in Availability possible support species This limited. a results ant protein carbohydrates. also These common are to colonies two relative with States. quantity within on increased individuals its United experiments growth and midwestern colony) independent provided the two ant amount and across the (i.e. of supported superorganism function was and a pattern as individual both where protein, pattern dietary robust a describe We DISCUSSION statistically ( highly carbohydrates again of was levels trials increasing with these increasing lab in compared preference The diet that the of visitation. P with double of effect carbohydrates, was level The more intermediate protein for an data. of received amount field protein least lower the ( of for significant the with level preference consistent with intermediate strong gradient were present The a the results ants had composite protein. to ants of a most the response number was the constant their ML the to held because for were protein; or preference especially carbohydrates of (carbohydrates) the When proteins, levels that numerator and to non-linear. hypothesized macronutrients was carbohydrates the We ratios both for by varied. in with preference driven one baits combined only was new a where offered ML of series we for in ratio, yet preference the colonies, the in (protein) whether shown). denominator not understand the (data to similar qualitatively order was low In result pair-wise the the of although and colonies, because 8), lab (P study (Fig. years) L this ( over diets in model preferred bait remaining significantly mixed-effects “L” and the were (termed intervals MH among neoniger content time and L. differentiate of protein several ML lack not that highest but a revealed the did field, or comparisons with species but the other diet content) in by the intrusion carbohydrate species avoid to each due to for discarded tended conducted be to were had trials stations baiting were 20 larvae About of needs carbohydrate the preference growth, Diet for workers to relative need protein high substantial. a if despite However, Thus, data. 5.00 the vs. in 0.18 cent evident significant, per not cent atomic statistically was per for being this atomic mean Larval but the likely. carbon, while more times to appears relative opposite larvae the to anything nitrogen more of movement for for collections 96-hour of enrichment .0)(i.S1). 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Data may be preliminary. u eeso uretpoiinn eeaporaet sesntin iiaina ucino ohthe both of function a as limitation nutrient those Therefore, diets. assess turn growth. provided to decreased to of for had week appropriate ratio ability however, rate P:C were per their weeks, growth and two provisioning once maximum saturated every amount nutrient a essentially fed once of availability found only Colonies levels we fed food Colonies that our Doubling provided. suggesting ants. weekly, food new diets. twice into of provided nutrients fed amount those the the than on better to colonies any respect grow with not did non-linear 2012). was 2009, growth Simpson, Colony & than Dussutour in rather than 2010; mortality 1:5 less al., measured and (but studies 1:3 et other between those (Cook lowest though the was diets, on mortality similar growth - sufficient using for reproduction between studies protein previous trade-off enough with a consistent and through both the are hand, achieved In and one is data toxicity). in the growth preliminary cause found on our optimal will was Together, storage that protein lipid effect. of suggest toxicity for studies levels paper a carbohydrates higher to this increased to even compared due in both with been presented with growth high have colony data & correlated relatively may in generally which Simpson all decrease study, was 2012; were A preliminary study our Roberts, study size. our con- worker & in our and protein content Woods, in number higher protein worker Harrison, ratios increased a 2012; P:C – toward toxicity 2009, the ratios, documenting However, Simpson, macronutrient & the 2009). 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Data may be preliminary. apig ilhl ietnl oeo h ope neatoswti ooista r icl oinfer to difficult are of that resolution colonies higher within a interactions and complex time-points the more of with some but study here, disentangle studies our employed Additional of help timeframe strategy assimilation. both will body). the pulse diet of their - sampling, in isotope hungry in/on isotopes castes were basic nitrogen larvae developmental labelled the until between labelled of workers using differences had in excess examine stored still to atomic being they insufficient showed were but was (i.e., proteins workers, still that diets than days possible unenriched carbon four is pattern fed of It at opposite excess colonies days. larvae an the to and of four was compared more workers either elements in significant), slightly for Both statistically gone having larvae larvae marginally was and nitrogen. with workers is workers not predicted transferred between result than in differences were (the transfer enrichment no nutrients nutrient anything were isotope of case, if there excess days the there, of four and necessarily after nutrient, signature is expectation, the As our but to optimal days. Contrary larvae, and four preferred to most over workers the colonies from of labelling through isotope carbohydrates) stable for Using workers. for and of carbohydrates true diet to be compared would disproportionately, opposite protein, the incoming that receive would preferences larvae that colony-level predicted We and colonies and eusociality within efficiency. Thus, flows metabolic constantly 2010). Nutrient increased scales Gillooly, to metabolism & regard scales basis, with rate Zanden, this advantageous mass Vander metabolic selectively of how per Kaspari, are extension with a (Hou, size line logical On this insects colony in A and superorganisms. increased is social insects and hypothesis 2010). and solitary organisms This Suarez, solitary than insect living. & The across limited in social Smith protein size content. to R less body benefit lipid is Chris inherent with development high 2008; an and their ratio) al., been growth C:N to have et insect lower may due much Smith social (i.e., largely R that content is is C nitrogen prediction this higher 2012; a workers, al., have have than et males, larvae (Schmidt ratio and While majority gynes (C:N) the workers. reproductive on nitrogen non-reproducing germline, premised non-growing, to colony’s is being prediction carbon superorganism This higher maximized a insects. is of a solitary growth studies biomass most superorganism standing these to that the compared in hypothesize of diet we that carbohydrate taxa, higher clear insect more a across is at a limited increasing currently It with with are protein). (maximized data better increased 2018). longevity While at as performed Long, such (higher flies ratios, & egg-laying P:C whereas and Buckiewicz, different major protein) max- 2009), Young, in two decreasing inherent to Behmer 2008; trade-offs these tend in life-history al., for grasshoppers, are (reviewed et choices there instar 1:1 nutritional (Lee last envi- above, their near and bias the noted to caterpillars P:C in carbohydrate As regard including protein with with 6). insects, of growth organisms Solitary quantity (Fig. imize by the carbohydrates 9). by faced of regulated (Fig. trade-off amount macronutrients was the a mass) by of thus lean regulated definition was some was or evolutionary there in content width the lipid seen head fit individual as as likely along ronment, (such supercolonies, not size, size that do its worker that 2002), note While expect Case, regulated; also & nutritionally would Tsutsui, is we Suarez, parts, then ‘organism’. Lach, constituent (Holway, super-superorganism its a species of existed ant processes there size conserved environment through nutritional If the achieved the is with with level. Kro- growth scales & cellular into size (Trible nutrients the variation that of size at is translation ‘organismality’ by the of explained because feature largely development universal is during a determination Thus, caste 2006; 2017). and Evans been nauer, 2007), & has Buck Robinson, it & (Wheeler, study, Toth signalling this 2008; Tor by in morphologically and expression and insulin behaviourally gene in regulated examine involved Patel is those not insects carbohydrates) including did social to we genes, some (relative nutrient-sensing While in protein determination provided. caste increasing diet that with of established increased amount size total ant the average and that demonstrated study Our Phenotypes Individual and Colony tal. et .neoniger L. 07 Toth 2007; e saple etae irgn(sapoyfrpoen n abn(saproxy a (as carbon and protein) for proxy a (as nitrogen traced we pulse, a as fed , tal. et 07 n eiwdi hnr ta. 08 mt,Tt,Sae,&Robinson, & Suarez, Toth, Smith, 2018; al., et Chandra in reviewed and 2007; 9 Posted on Authorea 21 Jan 2020 — CC BY 4.0 — https://doi.org/10.22541/au.157963861.14279623 — This a preprint and has not been peer reviewed. Data may be preliminary. G R n Saaye h aa GadCSwoetemnsrp,wihwsrvsdadapproved and revised was which manuscript, the wrote CRS and and YG care data. ant the through analysed study S. CS this Nicholson, and with E. CRS helped YG, Neumeister, all D. Zenker contributions: Lopez, M. Author Barrera and I. Serna T. Bailey, A. and M. Slavinskas, collection. Knorr Additionally, data K-H. S. Gadau, Saleh. Perkins, for J. data M. E. growth to Pant, and colony grateful Czaczkes Preliminary Crawley T. are YG. and B. We stoichiometry, to by and materials maintenance. nutrition providing and insect kindly housing on for colony discussions Schmidt on fruitful for insights Ott valuable D. for thank to like would We elucidated. ACKNOWLEDGEMENTS be multiple to assessing beginning by only diet are the and feedbacks, on information including decisions nutritional decisions, make integrate growth we to superorganisms into and which appear nutrition superorganisms. preferences, with by ants dietary translate and trade-off mechanisms of and also organisms The tuning – efficiency, for independently. both preference metabolic select ingredients of dietary to and of longevity, size predicted efficiency, are evidence the factors task found limit These as to such storage. resource up size, physiological of scales benefits limitation selective protein The that shown have stores/trash We to Cassill, or L 1976), (Deby Wilson, members 2009). O. nest Conclusion Simpson, E. across & 1985; distributed Vinson, (Dussutour ants are & protein nutrients excess, organisms Soren¸sen, Busch, with of the non-social 2005; enriched excretion and Wheeler, in and stomach & Although nutrients as protein Vinson, of colony. a not Butler, absorption the as selective is to larvae Furthermore, by it return the more mainly feedback. Therefore, use foragers composed to by be once resources regulated 2009). would nutrients not of mechanism Simpson, filtering is collection this & of foragers, the Dussutour through capable adjust expressed are 1999; and as ants Tschinkel, nest preference, levels. & the colony 2007). (Cassill in overall resource Tschinkel, larvae though presence low & of (Seal their studies of presence productivity/fitness in and with will the period 1984), protein preference to (Pyke, colonies current forager constraints responsive link of starved, the tightly are they types to survive Foragers when many failed because to have they example, ants due presumably should some optimal For 2007), on be being reproduction prioritizes not (Smith, That long-lived. may at this reproduction though, attempts be because Preference/foraging, conditions. in future to strategy investment laboratory have over adaptive tend under growth will an colonies only and in be growth assay, ant investment may from preference and prioritize foragers diet diet/bait growth, by (one-season) in over optimality preference single survival judge a carbohydrate to from increased conclusions difficult said, S1). generalizable also Fig. first draw is 8, the to it (Fig. in difficult field carbohydrates and of is lab levels It decreasing the higher in for the consistent at and was preference carbohydrates and in increasing diets decrease the for were on a of carbohydrates preference experiment caused each and preference clear for What protein preferences each a protein. clear When had of of ratio. amounts result ants the a the judging is individually, are diet the ants with manipulated the only the with in thus done carbohydrates alignment work and Further and macronutrients, in varying protein colonies). for not lab preference and was field that given between found preference consistent When colony were (data diet. growth, growth the colony colony in In assay protein preferred. in to were of discussed diets growth levels used As colony intermediate diets for metabolism. at same their optimum maximized fuel the is to growth of carbohydrates colony choices on that how primarily show and feed we nest, thus above, 2018). a and al., within non-growing, et sub-groups are (Shik different Workers assimilated to and nutrients of distributed distribution are relative they the rapidly as such behaviour, from .neoniger L. R n Gdsge h xeiet G P A n Mcletdtedata. the collected HM and AAW TP, YG, experiment. the designed YG and CRS .neoniger L. .niger L. nteohrhn,cln rfrnewsucniyaindwt optimal with aligned uncannily was preference colony hand, other the on , sucer u h ffc esse cosmlil rprtoso the of preparations multiple across persisted effect the but unclear, is ihrpoendesicesdgot,bthge carbohydrate higher but growth, increased diets protein higher – 10 .neoniger L. a collected was .neoniger L. Posted on Authorea 21 Jan 2020 — CC BY 4.0 — https://doi.org/10.22541/au.157963861.14279623 — This a preprint and has not been peer reviewed. Data may be preliminary. arsn .F,Wos .A,&Rbrs .P 21) urto,Got,adSz.In functions. Size. graphical and and Growth, statistical Nutrition, basic 9–62. diverse , RVAideMemoire: (2012). P. (2014). S. M. Herve, Roberts, & insects A., of H. physiology Woods, environmental F., J. Harrison, Ameisenstaat. “im (2018). J. ,,Soldaten M. Fox, der Entstehung . 808. . Die . D., (1937). 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YG to support Financial Funding authors. the of all by rceig fteRylSceyo odnB ilgclSciences Biological B: London of Society Royal the of Proceedings .niger L. netsSociaux Insectes , 12 olcinwsgatdb h iyo M of city the by granted was collection l a aawudb aepbil vial nteDydrepository. Dryad the in available publicly made be would data raw all l plcbentoa udlnsfrtecr n s faiaswr olwd Per- followed. were animals of use and care the for guidelines national applicable All netsSociaux Insectes 3,307–328. (3), h uhr elr htte aen optn interests. competing no have they that declare authors The nRCmaint ple Regression Applied to Companion R An ora fEprmna Biology Experimental of Journal , Vl ) xodUiest Press. University Oxford 3). (Vol. 55 , 3,329–333. (3), 49 4,320–325. (4), 11 nmlBehaviour Animal netsSociaux Insectes ntr(72-02 2017). (67-20-0032, unster ¨ nulRve fEntomology of Review Annual , Cell 219 urn Biology Current Naturwissenschaften , rspb20120051. , , 6,824–833. (6), aePublications. 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(1), netsSociaux Insectes . WNro Company. & Norton WW . rceig fteNtoa cdm fSciences of Academy National the of Proceedings nulRve fEooyadSystematics and Ecology of Review Annual S:fihre tc nlss akg eso ..20.9000 0.8. version package R analysis. stock fisheries FSA: ra nso ot mrc n uoe dnicto,booy n management and biology, identification, Europe: and America North of ants Urban , 21 ucinlEcology Functional 1,71–86. (1), , 41 7,545–553. (7), , 105 orsmn n vlto nisc societies insect in evolution and Nourishment eairlEooyadSociobiology and Ecology Behavioral netsSociaux Insectes 7,2498–2503. (7), h ueogns:tebat,eeac,adsrneeso insect of strangeness and elegance, beauty, the superorganism: The , 23 1,4–16. (1), akg Version Package R eairlEooyadSociobiology and Ecology Behavioral vial thTp/CA.RPoet Org/Package= R-Project. Ttp://CRAN. h at Available 12 , 59 , 2,289–295. (2), akg Version Package R 33 nulRve fEooyadSystematics and Ecology of Review Annual 200908071. , ymclgclNews Myrmecological 1,181–233. (1), , 1 (5). , 61 Ecology 8,1151–1160. (8), etiwpesBoulder, press Westview . lSOne PloS 1. , . gn Abn NY) (Albany Aging , 99 , , 26 59 9,1999–2009. (9), 81–96. , 2,215–221. (2), , 2 rceig of Proceedings 6,e509. (6), , 1 . , Posted on Authorea 21 Jan 2020 — CC BY 4.0 — https://doi.org/10.22541/au.157963861.14279623 — This a preprint and has not been peer reviewed. Data may be preliminary. hee,D . uk . vn,J .(06.Epeso fislnptwygnsdrn h eidof period the during genes Evolving pathway . insulin of (2006). mellifera. (2018). Expression J. H. Apis Wickham, (2006). bee, S. D. honey J. Simpson, the Evans, in & & determination N., D., caste Buck, Raubenheimer, E., P., D. Wheeler, K. insect. an Lee, in Pogonomyrmex T., obesity ant, to S. harvester resistance season. 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URL rceig fteNtoa cdm fSciences of Academy National the of Proceedings , eairlEooyadSociobiology and Ecology Behavioral 220 aueRvesGenetics Reviews Nature ora fIsc Science Insect of Journal 1,53–62. (1), 13 odn xodUiest Press. University Oxford London: . netMlclrBiology Molecular Insect , , 61 , , 9 17 6 1) 735. (10), 9,1479–1487. (9), (1). netsSociaux Insectes 3,191–198. (3), . , 33 h mrcnNaturalist American The 4,209–223. (4), , 15 , 103 5,597–602. (5), , 45 rnsi Genetics in Trends noo.News Entomol. 3) 14045–14049. (38), 4,385–410. (4), odSpring Cold odSpring Cold Functional , 172 , (4), 53 , Posted on Authorea 21 Jan 2020 — CC BY 4.0 — https://doi.org/10.22541/au.157963861.14279623 — This a preprint and has not been peer reviewed. Data may be preliminary. o ieetsca ridvda ris(otaeo)adteposo h ih hwdffrn piaand optima different show right the on plots the and optima overlapping trade-off) show (no left lines). the traits (intersecting on individual trade-offs plots thus The or social (bottom). colonies different within for individuals and (top) colonies social 1. Fig animals. in trade-offs history in life consequences of fitness physiology The and (2001). geometry Systematics G. Nutritional and L. Ecology Harshman, of (2018). & J., F. A. A. Zera, T. Drosophila. Long, Spotted-Wing the & species. suzukii, N., Drosophila ant colonies. Buckiewicz, an insect Y., of in Young, sociogenesis castes The of (1985). number O. the E. Wilson, and discretization Behavioral Sociobiology and (1976). Ecology O. E. Wilson, (1955). superorganism. O. the E. Reviving Wilson, Effect (1989). (2015). E. V. A. Sober, Suarez, & insect. 337–356. & social S., A., a D. D. in Holway, Wilson, condition D., and M. size, number, Eubanks, offspring M., One into S. investment PloS Wilder, on supplementation D., carbohydrate C. of Chong, D., B. Wills, (2016). Hadley. Wickham, (2010). Hadley. Wickham, ocpulshmtco h eainhpbtenntin viaiiyueadtatvlefrboth for value trait and availability/use nutrient between relationship the of schematic conceptual A , 10 7,e0132440. (7), oorpi eiino h n eu Lasius genus ant the of revision monographic A , 1 glt:eeatgahc o aaanalysis 1.4 data for v. graphics data elegant combining ggplot2: and applying splitting, for Tools plyr: , 2,141–154. (2), 32 1,95–126. (1), clg n Evolution and Ecology 14 Science , ora fTertclBiology Theoretical of Journal 228 Springer. . Vl 1) h Museum. The 113). (Vol. 40) 1489–1495. (4707), , 8 5,2842–2851. (5), . nulReview Annual Behavioral , 136 (3), Posted on Authorea 21 Jan 2020 — CC BY 4.0 — https://doi.org/10.22541/au.157963861.14279623 — This a preprint and has not been peer reviewed. 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