BIOLOGY Selected Life-History Traits of Black Soldier (Diptera: ) Reared on Three Artificial Diets

1 JEFFERY K. TOMBERLIN, D. CRAIG SHEPPARD, AND JOHN A. JOYCE

Department of Entomology, University of Georgia, Tifton, GA 31793

Ann. Entomol. Soc. Am. 95(3): 379Ð386 (2002) ABSTRACT illucens (L.) was reared on three larval diets to determine their effects on preimaginal development and selected adult life-history traits. Prepupal and adult characteristics were examined for individuals reared on each diet and compared with Þeld-collected prepupae and corresponding emergent adults. Diet did not signiÞcantly inßuence development or survivorship to the prepupal stage. However, adult emergence for all diets was signiÞcantly less than that determined for the wild population. Development time from egg to adult for individuals reared on the diets at 27ЊC ranged from 40 to 43 d with the larval stage lasting 22Ð24 d. We observed Ͼ96% larval survivorship to the prepupal stage and 21Ð27% adult emergence. Females accounted for 55Ð60% of emergent adults across treatments. Specimens reared on each diet were reduced in size, longevity, and calorie content in comparison to specimens from the wild population. Males within diet treatments and Þeld-collected specimens were signiÞcantly smaller than females and emerged 1Ð2 d before females. Additionally, males reared on the diets and provided water lived for 9 d, whereas females lived for 8 d. This information indicates the diets might be used for rearing soldier ßies. However, further reÞnement is needed to produce adults similar to those found in nature.

KEY WORDS Diptera, Stratiomyidae,

THE BLACK SOLDIER , Hermetia illucens (L.), is a large 1981) and swine (Newton et al. 1977). Prepupae, (13 to 20 mm), wasp-like ßy (May 1961). It has three when dried, have an estimated value comparable to generations per year in the southeastern United States menhaden Þsh meal. If used live, as specialty feed, or and can be collected from late spring through early fall marketed to exploit its other unique qualities (i.e., (Sheppard et al. 1994). Larvae occur in assorted de- essential fatty acids and chitin), the value of the prod- composing materials, such as fruits, , and ma- uct might be higher (Sheppard et al. 1994). Addition- nure (James 1935). ally, a system has already been developed for self- The black soldier ßy is interesting because its pres- harvesting the larvae by directing their search for ence can be used to solve many of the problems as- pupation sites into collection bins (Sheppard et al. sociated with large manure accumulations at conÞned 1994). feeding operations. House ßy, Musca domestica This manure management system depends on a ro- L., control due to manure being colonized by the black bust soldier ßy population for dependable inoculation soldier ßy has been reported by Furman et al. (1959), of the manure with larvae. Presently, little is known Tingle et al. (1975), Sheppard (1983), and Axtell and about the biology of H. illucens. Rearing of the black Arends (1990). Additionally, Tingle et al. (1975) doc- soldier ßy has been attempted in the past (Tingle et al. umented that black soldier ßy larvae reduce waste 1975) but was unsuccessful until 2002 (Sheppard et al. within poultry facilities. Sheppard et al. (1994) fol- 2002). With the development of suitable methods, the lowed up their work and determined that the black soldier ßy can now be produced and made available as soldier ßy can reduce manure accumulation by 42Ð a biological control and waste management agent. 56%. Concentrations of nitrogen, and other nutrients The primary objective of this study was to deter- are also signiÞcantly lower in this reduced manure mine basic biological parameters for the black soldier (D.C.S. unpublished data), thus further reducing the ßy when reared in selected diets. Additionally, we potential for pollution. compared life-history data for prepupae and adults Soldier ßy prepupae can be used as feed for a variety reared in these diets to data recorded for Þeld-col- of animals, including Þsh (Bondari and Sheppard lected prepupae and the emergent adults. Information from this study is important for improving rearing 1 Current address: Department of Biological and Agricultural En- methods necessary for integrating the black soldier ßy gineering, University of Georgia, P.O. Box 748, Tifton, GA 31793 into current waste management strategies in livestock (e-mail: [email protected]). and poultry facilities.

0013-8746/02/0379Ð0386$02.00/0 ᭧ 2002 Entomological Society of America 380 ANNALS OF THE ENTOMOLOGICAL SOCIETY OF AMERICA Vol. 95, no. 3

Table 1. Composition (%) of diets used to rear black soldier ture levels of 70%, whereas the CSMA diet moisture flies was slightly lower at 64%. For each diet during an experiment, 300 4-d-old soldier ßy larvae were placed Gainesville Layer hen Constituent CSMA diet ration in each of three 1-liter clear plastic cups. Cups were each covered with a paper towel and held in the Alfalfa meal 30.0 27.0 a Wheat bran 50.0 33.0 a rearing room. Feeding was terminated in a treatment Corn meal 20.0 Ñ a when a cumulative 40% of the larvae in the three cups BrewersÕ dried grain Ñ 40.0 Ñ reached the prepupal stage. However, daily observa- Factors tions continued until all larvae had entered the pre- Protein 15.3 19.0 15.0 Fat 3.8 3.0 3.0 pupal stage or died. Prepupae were identiÞed by a Fiber 12.6 20.0 5.0 change in integument color from larval white to black Ash 6.3 8.0 13.7 (May 1961). Calcium 4.9 4.0 5.1 Immature Life-History Traits. Larval weight in a Proprietary information, but comparable to that placed in other each treatment was determined daily by selecting 10 treatments. larvae from each cup per treatment and weighing them individually on a Mettler AT261DeltaRange balance (Mettler-Toledo, Hightstown, NJ). After Materials and Methods weighing, larvae were returned to their respective cups. Final larval weight was deÞned as the average Acquisition of Flies. At a 100 bird caged-layer poul- larval weight recorded on the day when 40% of all try house at the Coastal Plain Experiment Station in individuals in a treatment reached the prepupal stage. Tifton, GA, we collected 10Ð20 female soldier ßies Prepupae were removed daily and transferred to probing the manure surface with their ovipositors and another 1-liter cup (containing 100 g white sand as a placed them in a 0.5-liter glass jar witha2by3-cm roll pupation medium) designated to hold prepupae from of corrugated cardboard as an oviposition substrate that particular replicate of a diet treatment. Addition- (Booth and Sheppard 1984). After 24 h, the cardboard ally, each day, three of the prepupae collected from containing soldier ßy eggs was placed in another 0.5- each replicate of each treatment were weighed and liter Sweetheart plastic cup (Sweetheart Cup Com- placed in petri plates and stored at Ϫ15ЊC for later pany, Chicago, IL) covered with a dry paper towel, determination of caloric content. Cups containing and stored in a rearing room (27ЊC, 60Ð70% RH, and prepupae were covered with 50 by 50 cm of mosquito a photoperiod of 14:10 [L:D] h) until larvae emerged netting held in place with a rubber band, placed in the 4 d later. Once larvae were observed, 50 g of a 15% rearing chamber, and monitored daily for adult emer- protein laying hen ration (Country Acres Feed, Brent- gence. Percentage of larvae reaching the adult stage wood, MO) mixed with 90 ml of water (70% moisture) was placed in the container. The larvae were allowed was recorded for each diet treatment. This could not to feed on the medium for 4 d before being used in the be determined for the wild population, which was experiment. sampled when ßies reached the prepupal stage. Days Wild Fly Population. Black soldier ßy prepupae from egg collection to adult emergence were recorded were collected from swine and poultry units located in for each ßy reared on the diets. Tifton, GA, and held in the above mentioned rearing Prepupal Calorie Content. Mean caloric content of room in six 1-liter clear Sweetheart plastic cups at a prepupae per treatment was determined using bomb density of Ϸ500 per cup. Prepupal and adult charac- calorimetry (combustion) techniques (Dutcher 1983). teristics compared across diet treatments were com- Frozen prepupae were held from seven to 14 d in a Њ pared with data recorded for the wild population (see convection oven set at 55 C. Individual dried prepu- below). pae were then weighed on a Mettler EA 100 electronic Ϸ Experimental Design. The following three diets balance (Metler-Toledo), ground to 1-mm pieces, were evaluated (Table 1): (1) Gainesville diet, which mixed with calorimeter grade benzoic acid (Sigma, was developed for rearing house ßies (Hogsette 1985) St. Louis, MO) (1:10 specimen weight: benzoic acid and the diet currently suggested for mass rearing the weight) and compressed into a pellet. We were unable black soldier ßy (Sheppard et al. 2002); (2) CSMA, an to make pellets with prepupae collected from the wild. artiÞcial rearing media for house ßy larvae (Chemical These prepupae, unlike those reared on the diet treat- Specialties ManufacturersÕ Association, Ralston Pu- ments, appeared to contain greater amounts of fats and rina, St. Louis, MO); (3) the 15% protein layer hen attempts to crush oven-dried specimens resulted in ration, which had been used in preliminary trials for observable loss of materials. Therefore wild prepupae rearing H. illucens. were analyzed without being mixed with benzoic acid. The experiment was replicated three times from To ensure that this difference in techniques did not April through October 1999. Preliminary studies using inßuence the results, caloric content was determined the layer hen ration rate of 10 g mixed with 17 ml water for prepupae from each diet treatment prepared with to 300 larvae daily resulted in the heaviest Þnal larval and without benzoic acid. A Parr oxygen bomb calo- weight. Therefore, this rate was selected as the feeding rimeter (model No. 1341) (Parr Instrument, Moline, rate to be used for all three diets. The layer hen ration IL) was used to determine caloric content of pellets. and Gainesville diet mixed with water had initial mois- Caloric content per mg ash-free dry weight per pre- May 2002 TOMBERLIN ET AL.: SELECTED LIFE-HISTORY TRAITS FOR BLACK SOLDIER FLIES 381 pupa was determined for each diet treatment and for 5-liter bucket (Plastic Packaging, West SpringÞeld, the wild population. MA) containing Ϸ1kg Gainesville diet saturated with Adult Life-History Traits. Weight, sex, and time water was placed on a cinder block (40 cm high) in from oviposition to adult emergence were recorded each cage to attract ovipositing individuals. Methods for each adult from the diet treatments. Individual for collecting eggs were adapted from Booth and adults were placed in 35-ml Solo cups (Solo, Urbana, Sheppard (1984). Three layers of corrugated card- IL) capped with a breathable lid. Half of the adult ßies board (each ßute opening measured 1by 4 mm) were were provided 0.125 ml water daily via a 1.25-cm glued together, cut into 5.0 by 2.5 by 1.0-cm blocks, needle inserted through the lid. The remaining ßies and taped inside the 5-liter bucket Ϸ3 cm above the were held in similar conditions, but without water. media. These cardboard blocks were replaced daily. Longevity was recorded for both groups. An egg clutch was deÞned as a ßute containing Ͼ100 Egg Development and Clutch Size. Initially, we eggs. Egg clutches collected each day from each treat- hypothesized that larval media inßuences the number ment were weighed and stored in vials containing 80% of eggs produced by black soldier ßies and that mating ethanol until the eggs could be counted. was not required to produce eggs. Therefore, to test Egg Collection System. We suspected that eggs per this hypothesis and determine the number of eggs per ßute did not accurately represent the number of eggs individual, a sample of nonmated females from each oviposited per individual. Therefore, we conducted an diet treatment and from the wild population was dis- experiment to evaluate the relationship between ßute sected in saline solution in a petri plate 1Ð4 d after dimensions and eggs deposited per ßute by black sol- emergence. Ovaries were removed and the number of dier ßies reared in our colony and held ina2by2by developed eggs recorded. Specimens were dissected 3-m cage placed in the greenhouse. We examined and examined under a Leica2000 Zoom dissecting mi- three cardboard ßute sizes; one with a smaller opening croscope (Leica, Buffalo, NY). A fully developed egg (1by 3 mm); one with the same opening (1by 4 mm); was deÞned as elongate and creamy to white in color, and the third with a larger opening (2 by 5 mm) than while an oocyte was deÞned as spherical and trans- that used in the diet experiments. Cardboard blocks parent. Oviposition by females in 35-ml cups was also were collected and replaced daily. Eggs from each recorded to determine if the number of eggs in dis- ßute were stored in a vial containing 80% ethanol. Egg sected specimens was similar to the number deposited number per ßute was determined as described above. by females reared on the same diet. Statistical Analysis. Procedure GLM (SAS Institute Dissected, unmated specimens did not contain de- 1992) was used to analyze the recorded data. Least veloped eggs. Therefore, we suspected that mating signiÞcant difference (LSD) test (SAS Institute 1992) was required for egg production. To test this hypoth- was used following a signiÞcant F test (P Յ 0.05) to esis, Ϸ500 (Ͻ24 h postemergence) black soldier ßies separate mean differences between diet treatments, were collected from a colony (larvae fed Gainesville and the wild population for: percent survival to adult diet) and released at 1500 hours ina2by2by3-mcage stage, percent females and males to emerge, pupal skin in a greenhouse maintained at Ϸ30ЊC. Observations weights, adult male and female weights, and male and and collection of mating pairs were made hourly be- female longevity with and without water. Additionally ginning at 1600 hours on the day the adults were for the diet treatments only, the previously described released and continued daily from 0800 to 1700 hours. procedure was used to separate mean larval and pupal Fifty-Þve mating pairs were collected by the conclu- development periods and Þnal larval weight. Percent- sion of the second day. Mated females were weighed age values were normalized by arcsine transformation. within 1h of mating, placed in capped 35-ml cups, and Mean egg number and total weight of eggs per ßute held in the rearing room and provided water as pre- were analyzed using the procedures previously de- viously described. We determined in a previous ex- scribed. Regression and PearsonÕs correlation analysis periment that oviposition generally occurred 2 d after (SAS Institute 1992) were used to determine the re- mating (Tomberlin 2001). Therefore, 2 d after mating, lationship between egg number and mass weight per the ovaries of 45 mated females were dissected and ßute for each treatment, and adult weight and lon- percent with developed eggs determined. Number of gevity. developed eggs per individual was also recorded. The remaining 10 females were allowed to oviposit Results in corrugated cardboard taped to the opening of the 35-ml cup. These eggs were removed and the number Immature Life-History Traits. Final larval weight determined based on mass weight (Booth and Shep- across treatments ranged from 0.153 to 0.171 g (Table pard 1984). The estimated number of eggs deposited 2) and did not differ signiÞcantly (F ϭ 2.35; df ϭ 2, 2; in the cardboard was compared with the number of P Ͼ 0.2118). Survivorship of larvae to the prepupal eggs recorded for dissected mated females. stage was not signiÞcantly different across diet treat- To determine the number of eggs oviposited by ments (F ϭ 1.07; df ϭ 2, 23; P Ͼ 0.3592) and ranged adults reared on each of the three diets and from the from 96.0 to 97.8% (Table 3). Time from egg to pre- wild population, we released Ϸ500, Ͻ24 h old, adults pupal stage per treatment ranged from 22.5 to 24.1d from each treatment into 2 by 2 by 3-m cages placed (Table 4) and did not differ signiÞcantly (F ϭ 0.74; in the greenhouse previously described. Each treat- df ϭ 4, 4; P Ͼ 0.6121). Mean prepupal weights differed ment had a separate cage. To collect egg masses, a signiÞcantly among diet treatments including the wild 382 ANNALS OF THE ENTOMOLOGICAL SOCIETY OF AMERICA Vol. 95, no. 3

Table 2. Comparison of larval, prepupal, pupal skin, and adult weights (grams), and prepupal caloric content per milligram of ash-free dry weight for H. illucens reared on three diets and from field-collected prepupae

Prepupal Pupal skin Adult weight Diet Final larval weight Caloric content/mg weight weight Male Female Gainesville diet 0.157 Ϯ 0.077A 0.104 Ϯ 0.027A 3.51 Ϯ 0.28A 0.011 Ϯ 0.003A 0.046 Ϯ 0.005A,a 0.056 Ϯ 0.005A,b n ϭ 3 n ϭ 30 n ϭ 23 n ϭ 40 n ϭ 272 n ϭ 339 CSMA 0.153 Ϯ 0.063A 0.107 Ϯ 0.041A 4.21 Ϯ 0.39AB 0.011 Ϯ 0.002A 0.044 Ϯ 0.005A,a 0.054 Ϯ 0.005A,b n ϭ 3 n ϭ 30 n ϭ 22 n ϭ 45 n ϭ 254 n ϭ 320 Layer hen ration 0.171 Ϯ 0.043A 0.111 Ϯ 0.034A 4.48 Ϯ 0.27B 0.016 Ϯ 0.006B 0.053 Ϯ 0.007B,a 0.064 Ϯ 0.006B,b n ϭ 3 n ϭ 30 n ϭ 20 n ϭ 45 n ϭ 234 n ϭ 316 Wild population NA 0.220 Ϯ 0.040B 5.95 Ϯ 0.32C 0.025 Ϯ 0.008C 0.085 Ϯ 0.016C,a 0.111 Ϯ 0.022C,b n ϭ 30 n ϭ 15 n ϭ 30 n ϭ 57 n ϭ 70

Means within a column followed by different capital letters are signiÞcantly different. Means for both sexes with different lowercase letters within a treatment differ signiÞcantly (P Յ 0.05; LSD, SAS Institute 1992). NA, Not applicable. population (F ϭ 146.79; df ϭ 3, 116; P Ͻ 0.0001) and soldier ßies reared on diets were signiÞcantly smaller ranged from 0.104 to 0.220 g (Table 2). Pupal skins than females (F ϭ 493.59; df ϭ 7, 1854; P Յ 0.0001) accounted for Ϸ10% of their weight (Table 2). Pre- reared on the same diet. This size differential also pupae reared on layer hen ration had the greatest occurred in the wild population (Table 2). However, numerical weight among diet treatments, but still when comparing the same sex across treatments, wild weighed signiÞcantly less than prepupae from the wild males and females were signiÞcantly larger than their populations. Prepupal skin weight differed signiÞ- diet-reared counterparts. Although not signiÞcantly cantly between diet treatments (F ϭ 75.47; df ϭ 3, 156; different (F ϭ 9.92; df ϭ 7, 10; P Ͼ 0.2875), adult males P Յ 0.0001) with skins from specimens reared on layer generally emerged earlier than females (Table 4). hen ration and from the wild population weighing the An interaction effect was determined for larval diet most. and the provision of water on adult longevity (F ϭ Prepupal Calorie Content. Mean calorie content 48.22; df ϭ 15, 1,764; P Յ 0.0001) of black soldier ßies per mg ash-free dry weight per prepupa was signiÞ- (Table 4). Adult males reared on a diet and provided cantly different across treatments (F ϭ 8.70; df ϭ 3, 76; water as adults lived 9.3Ð9.7 d, whereas those not P Յ 0.0001) and ranged from 3.51 to 5.95 cal per mg provided water lived 6.0Ð7.1d. Females reared on the ash-free dry weight, with those specimens collected treatments and provided water as adults lived 7.9Ð8.5 from the wild having the greatest calorie content (Ta- d, whereas those not provided water lived 6.1Ð6.4 d. ble 2). Wild prepupae weighed more and had a higher Males and females that emerged from wild-collected Ϸ concentration of fat, yielding 3.5 times more calo- prepupae lived signiÞcantly longer than those reared ries, than prepupae reared on the diet treatments. on the artiÞcial diets. Wild males and females lived Ϸ Prepupae reared on the diets contained 365 calories, Ϸ14 d when provided water as adults and 7.8 and 8.2 d, whereas the wild prepupae contained an average of respectively, when not provided water. Male and fe- 1,309 calories. No signiÞcant differences were deter- male longevity within a treatment when not provided mined for the caloric content of prepupae prepared water did not differ. Longevity of adults provided with or without benzoic acid. water was signiÞcantly correlated with individual Adult Life-History Traits. Mean adult emergence weight (r2 ϭ 0.40; P Յ 0.0001). rate (Table 3) differed signiÞcantly (F ϭ 26.84; df ϭ Ն Egg Development and Clutch Size. Twenty virgin 3, 27; P 0.0001) and ranged from 21.7 to 27.2% for the females per treatment, and four from the wild popu- diet treatments, whereas 91.3% of the prepupae sam- lation, from 1t o 4 d postemergence were dissected and pled from the wild yielded adults. Of those to emerge, examined for mature eggs, but none were found (Fig. 55.2Ð60.5% were female (Table 3) with no signiÞcant 1a and b). For each treatment, including the wild difference across diet treatments or the wild popula- Ͻ ϭ ϭ Ͼ population, 1% of the virgin females placed in plastic tion (F 1.05; df 2, 22; P 0.3660). Male black medicine cups laid eggs. None of these eggs hatched. Thirty-two (72%) of the 45 mated-colony females that Table 3. Percent survival of H. illucens larvae to the prepupal were dissected 2 d after mating (Fig. 1c) had com- and adult stages and sex ratio for larvae reared on three diets and pletely formed eggs. Number of eggs per individual for prepupae from a wild population ranged from 323Ð621, which accounted for Ϸ13Ð26% Larval Adult of their body weight. Six of the 10 mated females from Sex ratio Diet n survivorship emergence the colony that were held individually in 35-ml cups (% female) to prepupae from prepupae deposited eggs. The number of eggs laid per individual Gainesville diet 8 97.8 Ϯ 0.6A 27.2 Ϯ 6.7A 60.5 Ϯ 1.2A ranged from 206 to 639, which accounted for 7.9Ð CSMA 9 96.0 Ϯ 1.4A 23.9 Ϯ 4.0A 55.2 Ϯ 2.0A 23.4% of their body weight. Individuals dissected 3 d Ϯ Ϯ Ϯ Layer hen ration 9 96.1 0.4A 21.7 1.8A 58.0 3.4A after laying eggs (Fig. 1d) had no additional ovarian Wild population 6 NA 91.3 Ϯ 4.8B 56.0 Ϯ 0.0A development and little fat body present. ϭ Means within a column followed by different letters differ signif- Egg Collection System. Mean eggs per ßute (F icantly (P Յ 0.05; LSD, SAS Institute 1992). NA, not available. 0.72; df ϭ 3, 71; P Ͼ 0.5456) and weight of eggs per ßute May 2002 TOMBERLIN ET AL.: SELECTED LIFE-HISTORY TRAITS FOR BLACK SOLDIER FLIES 383

Table 4. Selected life-history traits of H. illucens reared on three diets and for adults reared from field-collected prepupae

Longevity, d adults Longevity, d adults Development, Development from Diet treatment provided water not provided water d from egg egg to adult, d Male Female Male Femaleto prepupa Male Female Gainesville diet A9.3 Ϯ 0.4A,a A7.9 Ϯ 0.2A,a B6.0 Ϯ 0.2A,a B6.1 Ϯ 0.1A,a 22.5 Ϯ 0.7A 43.0 Ϯ 2.9A,a 43.4 Ϯ 2.1A,a n ϭ 106 n ϭ 161 n ϭ 159 n ϭ 160 n ϭ 3 n ϭ 3 n ϭ 3 CSMA A9.7 Ϯ 0.4A,a A8.5 Ϯ 0.2A,a B5.9 Ϯ 0.2A,a B6.2 Ϯ 0.2A,a 23.4 Ϯ 0.3A 43.0 Ϯ 2.5A,a 43.0 Ϯ 1.3A,a n ϭ 103 n ϭ 163 n ϭ 145 n ϭ 138 n ϭ 3 n ϭ 3 n ϭ 3 Layer hen ration A9.3 Ϯ 0.4A,a A8.5 Ϯ 0.3A,a B7.1 Ϯ 0.2B,a B6.4 Ϯ 0.2A,a 24.1 Ϯ 0.9A 40.4 Ϯ 2.4A,a 41.7 Ϯ 2.1A,a n ϭ 96 n ϭ 151 n ϭ 140 n ϭ 138 n ϭ 3 n ϭ 3 n ϭ 3 Wild population A14.3 Ϯ 1.2B,a A14.2 Ϯ 0.9B,a B7.8 Ϯ 0.4B,a B8.2 Ϯ 0.4B,a NA NA NA n ϭ 25 n ϭ 42 n ϭ 31 n ϭ 22

Means within a column followed by different capital letters differ signiÞcantly. Means for both sexes within a treatment with different lower case letters differ signiÞcantly. Longevity means for a sex provided water and not provided water preceded by different capital letters differ signiÞcantly (P Յ 0.05; LSD, SAS Institute 1992). NA, Not available.

(F ϭ 0.83; df ϭ 19, 20; P Ͼ 0.4798) were not signiÞ- larvae fed 5-d-old hen manure grew at half the rate of cantly different among diet treatments, including the larvae fed 18 h old manure (D.C.S., unpublished data). wild population (Table 5). Individual eggs ranged in Weight differences recorded for prepupae reared on weight from 0.23 to 0.25 mg and mean number of eggs laying hen ration and those reared on the other diets per ßute ranged from 603 for adults reared on Gaines- might be attributable to differences in calcium con- ville diet and the wild population to 689 for adults tent (Table 1). Soldier ßy larvae sequester ingested reared on layer hen ration. Mean weight of eggs per calcium and convert it to calcium carbonate, which is ßute (clutch weight) ranged from 0.0145 g from adults secreted through the hypodermis and covers the larval reared on the Gainesville diet to 0.0159 g for adults integument (Johannsen 1922). reared on the CSMA larval ßy diet. Correlations be- Development time from egg to prepupae for the tween mean egg weight and number of eggs per ßute black soldier ßy in our study was six to eight days for adults reared on each diet and the wild population shorter than that recorded by May (1961) who re- were signiÞcant (P Ͻ 0.05) (Table 5). The regression ported a mean development of 31d under similar analysis for mean egg number versus egg clutch weight temperature and humidity conditions. Differences be- per ßute size was also signiÞcant for all treatments tween the two studies might be due to various factors, including the wild population (Table 5). such as different larval densities (Morrison and King The inability to determine a difference in the size of 1977) or food quality (Slansky and Scriber 1985, Dicke egg clutches per diet treatment and the wild popula- et al. 1989). Furman et al. (1959) suggested these tion was due to the early misconception that one ßute factors could delay black soldier ßy larval develop- held the eggs of one female (Booth and Sheppard ment up to four months. However, May (1961) did not 1984). The number of eggs deposited per ßute was provide detail sufÞcient for comparison. limited by ßute size. The number of eggs deposited in Differences in percent emergence per diet treat- the smaller ßute (1by 3 mm) averaged 431eggs per ment and those collected from the wild might also be ßute, whereas those in the larger ßute (2 by 5 mm) due to larval diet and being held in a sub-optimal averaged 1,157 eggs per ßute. Based on measurements habitat. Larvae in our experiment were held and fed of the depths that eggs were deposited in the different in containers that resulted in continuous contact with sized ßutes, individuals were able to insert their ab- their excreta. Sheppard (1983) determined that wild domen and ovipositor Ϸ1cm into the larger ßute and soldier ßy larvae remain on the surface of the wastes Ϸ0.7 cm in the smallest ßute. Egg size was not signif- accumulating below layer facilities and feed on fresh icantly different across diet or ßute treatments (F ϭ manure as it is produced. Manure digested by the 0.83; df ϭ 3, 71; P Ͼ 0.4798). larvae then accumulates below the larvae. Because the soldier ßy larvae in our experiment were held in closed containers, they fed on media that was mixed with diet Discussion residue and their own waste. Therefore, the soldier ßy All three diets are suitable for rearing the black larvae might have been forced to feed to a degree on soldier ßy. They are nutritionally similar, which prob- their own wastes, which we suspect they normally ably explains why Þnal larval weight and caloric con- avoid. Larvae of other dipteran species, such as the tent of diet-reared prepupae were comparable. How- house ßy, need fresh manure daily for optimal growth ever, lab-reared prepupal weight differed from the (Morgan and Eby 1975). This need for fresh manure wild prepupae sampled, which might be due to wild is because anaerobic organisms digest manure as it larvae having access to additional nutrients (Slansky ages, resulting in fewer nutrients for the larvae and and Scriber 1985, Dicke et al. 1989), such as the con- thereby suppressing larval growth (Beard and Sand tinuous stream of fresh manure in poultry facilities, 1973). Developmental time from egg to the adult did not mixed with old resources as in our tests. Recent not differ between sexes for wild black soldier ßies, as laboratory studies determined that black soldier ßy well as those reared on diet in the laboratory. 384 ANNALS OF THE ENTOMOLOGICAL SOCIETY OF AMERICA Vol. 95, no. 3

Fig. 1. Ventral view of the dissected abdomen of a female black soldier ßy. (a) Freshly emerged (Ͻ24 h) with fat body present; (b) freshly emerged (Ͻ24 h) with fat body removed; (c) 2 d after mating remaining fat body intact; (d) 3 d after laying eggs with remaining fat body intact.

Water, unlike food, is essential for adult black sol- adults being unable to mate effectively and therefore dier ßies to reproduce (Sheppard et al. 2002), which not laying viable eggs. We determined that individuals might be due to the less vigorous and dehydrated reared on the diets and provided water lived 1Ð2 d May 2002 TOMBERLIN ET AL.: SELECTED LIFE-HISTORY TRAITS FOR BLACK SOLDIER FLIES 385

Table 5. Comparison of clutch weight (grams), and egg weight (milligrams) for H. illucens reared on three diets and for field-collected prepupae and the correlation and regression analyses for clutch weight (milligrams) and eggs per clutch per treatment

Treatment Correlationa Regression models for eggs per clutch Clutch weight Egg weight Gainesville diet r2 ϭ 0.88, df ϭ 20 ϭ 173.87 ϩ 29,372 (clutch weight) 0.0145 Ϯ 0.00012A 0.24 Ϯ 0.012A n ϭ 20 F ϭ 62.8; df ϭ 1, 18; P Ͻ 0.0001 CSMA r2 ϭ 0.90, df ϭ 15 ϭ 143.15 ϩ 31935 (clutch weight) 0.0159 Ϯ 0.00018A 0.25 Ϯ 0.011A n ϭ 15 F ϭ 55.5; df ϭ 1, 13; P Ͻ 0.0001 Layer ration feed r2 ϭ 0.93, df ϭ 20 ϭ 124.64 ϩ 35346 (clutch weight) 0.0158 Ϯ 0.00012A 0.23 Ϯ 0.008A n ϭ 20 F ϭ 122.3; df ϭ 1, 18; P Ͻ 0.0001 Wild population r2 ϭ 0.98, df ϭ 20 ϭ 25.12 ϩ 38887 (clutch weight) 0.0153 Ϯ 0.00012A 0.25 Ϯ 0.004A n ϭ 20 F ϭ 686.8; df ϭ 1, 18; P Ͻ 0.0001

a PearsonÕs correlation, signiÞcance set at P Յ 0.05 (SAS Institute 1992). Means within a column followed by different letters differ signiÞcantly.

longer than those not provided water. In contrast, wild estimate of the total number of eggs produced by a adult soldier ßies provided water lived 4 d longer than female. Additionally, oviposited eggs might not be as adults reared on the diets. This signiÞcant difference accurate due to variables inßuencing number of eggs in longevity might be explained by caloric content of produced, such as time from emergence to mating. the prepupae. Prepupae reared on the diet treatments Rogers and Marti (1994) determined that the age of Ϸ contained 365 cal, whereas the wild prepupae con- female fall armyworms, Spodoptera frugiperda (J. E. tained 1,309 cal per prepupa. The additional calories Smith) (Lepidoptera: Noctuidae), at Þrst mating sig- provided energy that might have resulted in increased niÞcantly inßuences reproductive potential and that longevity of the wild adults. extended female virginity resulted in reduced fecun- Egg development studies were problematic because dity and fertility. of misconceptions concerning the deÞnition of a black The black soldier ßies in our studies reproduced soldier ßy egg clutch, or eggs oviposited by one indi- without feeding and apparently relied on fat reserves vidual. Booth and Sheppard (1984) stated that the average number of black soldier ßy eggs per clutch was acquired during larval development for adult sur- 998 Ϯ 78 (mean Ϯ SE). They collected eggs using the vival and egg production (D.C.S., unpublished data). openings in corrugated cardboard (ßutes) as the ovi- Therefore, any delay in mating could result in re- position site. Before Booth and Sheppard (1984), re- sources being reallocated from producing eggs to pro- searchers reported black soldier ßy egg clutches with longing the adult stage. Similar to our hypothesis for 119Ð502 eggs (Gonzalez et al. 1963) and 205Ð802 (Ste- the black soldier ßy, Chippindale et al. (1993) deter- phens 1975) in dried hen manure and on bananas, mined that food quality was directly related with egg respectively, which are similar to our eggs per clutch production and inversely related to adult longevity for data determined for individuals dissected 2 d after Drosophila melanogaster Meigen (Diptera: Drosophi- mating. lidae). Higher quality food resulted in greater egg Methods described by Booth and Sheppard (1984) production, but reduced longevity in fruit ßies. Ad- for collecting black soldier ßy eggs are suitable. How- ditionally, the number of times a female arthropod ever, these methods are not recommended for esti- mates can inßuence the number of eggs deposited. As mating clutch size for individual soldier ßies. Egg suggested by Foster and Ayers (1995) for female clutches from diet-reared and wild adults were smaller Epiphyas postvittana (Walker) (Lepidoptera: Tortri- than those recorded by Booth and Sheppard (1984), cidae), mating more than once might be attributed to but individual egg size was similar. The reduction in male deÞciencies, but might result in greater numbers egg number per clutch for our experiment and that of eggs being fertilized and deposited. recorded by Booth and Sheppard (1984) is most likely Most of the literature on the black soldier ßy is due to ßute size, which we determined can limit the restricted to its use as a biological control agent of number of eggs deposited per ßute. Therefore, our house ßies and as a manure management agent in results really estimate number of eggs per ßute, which poultry facilities. Our study provides additional infor- also might have been the case for Booth and Sheppard (1984). However, the ßute size they used is unknown. mation on the life history of this species reared on We suspect that multiple individuals could deposit three diets, as well as for those collected from the wild. eggs in a single ßute, which would skew clutch size Such information is necessary for developing its po- estimates. Egg clutch weight varied Ͻ1% when allow- tential as a biological control and waste management ing individuals to oviposit in cardboard ßutes of equal agent in livestock and poultry production. The three size. However, egg clutches oviposited by mated in- diets examined in this study are suitable for rearing dividuals collected from the black soldier ßy colony black soldier ßies. However, because of the large dif- and placed in 35-ml cups and those dissected 2 d after ferences between wild and laboratory-reared speci- mating varied as much as 75%. We hypothesize dis- mens, further research is needed to reÞne and improve secting females 2 d after mating provides an accurate larval rearing for mass production of this species. 386 ANNALS OF THE ENTOMOLOGICAL SOCIETY OF AMERICA Vol. 95, no. 3

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