Larval Substrate Research Articles

Immunity-related genes can serve as early diagnostic markers for infection in housefly rearing systems

Abstract Farming of houseflies, as well as of other insects, could become a sustainable alternative for producing high-protein animal feed. Although promising, this comes with particular challenges, including monitoring insect health in large-scale farming facilities. In this study, we tested if the expression patterns of immunity-related genes could serve as early diagnostic markers for disease and help prevent infection spread with detrimental effects. We introduced a commonly used bacterial entomopathogen, Bacillus thuringiensis, in the housefly larval substrate and monitored larval biomass changes, pupation, and gene expression of three immunity-related genes: PGRP, defensin, and cecropin. We performed the infection assay for two strains of different geographic origins at two temperatures and with two concentrations of the bacterial inoculum. We found that gene expression was faster in indicating differences between larvae grown in treated and untreated substrate compared to monitoring larval growth and pupation. Most strikingly, although there were no significant effects on larval biomass for 48 hours after the substrate inoculation, the effects on gene expression in some cases manifested already two hours post substrate inoculation. As the two housefly strains showed different response speeds in gene upregulation, there is a need for more frequent routine testing to define which response levels could be considered as an alarm. We conclude that monitoring immunity-related genes and microbial surveys of insect-rearing substrates can be used to diagnose infection and prevent its spread early in mass-rearing settings.

Use of different dry materials to control the moisture in a black soldier fly (Hermetia illucens) rearing substrate.

Controlling the substrate moisture is a significant challenge in black soldier fly (BSF) farming. Many substrates have a high moisture content, which results in a low BSF biomass and a high mortality. One potential solution involves incorporating dry substrates into the food mix to mitigate the excessive moisture. However, little information about the types and quantities of dry substrates is available. Six different dry materials-rice husk (RH), rice bran (RB), rice husk ash (RHA), coconut coir dust (CC), rubberwood sawdust (RSD), and spent coffee grounds (SCGs)-were evaluated by combining with pure minced mixed vegetables in varying proportions (0%, 5%, 10%, 15%, 25%, and 50% by weight). This study encompassed both small-scale and medium-scale experiments to comprehensively assess the effects of the addition of each of these different dry substrates and their quantities on aspects of the development of BSF, such as BSF biomass, larval duration, mortality rates, adult sex ratio, and the moisture removal efficiency of each substrate mixture. Each dry substrate had specific properties. Although RB emerged as a favorable dry substrate owing to its nutritional content and substantial water-holding capacity, excessive use of RB (>15% by weight) resulted in elevated temperatures and subsequent desiccation of the substrate, potentially leading to larval mortality. In contrast, RH demonstrated the ability to support improved larval duration and growth, permitting its utilization in higher proportions (up to 50%). On the other hand, CC, RHA, and SCG are better suited for inclusion in BSF larval substrates in smaller quantities. Some dry substrates require a pretreatment process to eliminate toxic substances prior to their incorporation into substrate mixtures, such as CC and SCG. A potential alternative solution involves employing a combination of various dry substrates. This approach aims to enhance the substrate moisture control and subsequently improve the BSF rearing performance.

Effect of two larval diets on the reproductive parameters of the housefly, Musca domestica L. (Diptera, Muscidae)

Abstract Deep understanding of the optimal reproductive and biological parameters is necessary to obtain the maximum egg production in captive rearing of Musca domestica, the common housefly. The effect of larval diet is explored in this study, using two different media: poultry manure and an artificial medium based on cereals mixed with meat. The preimaginal mortality and development time, with other reproductive parameters that indirectly affect adult fecundity as: ovary and wing size, fluctuating asymmetry, and cuticular hydrocarbon composition, were analysed. The results show that poultry manure was related with better efficacy values, lower larval mortality rate, and a sex-dependent effect in fecundity. Asymmetry fluctuation and different hydrocarbon composition was detected in females. The females from larvae that grew on manure presented bigger ovaries and oocytes, but a smaller number of ovarioles per ovary. Their morphological asymmetry was also lower; and the cuticular hydrocarbon composition was distinct from the rest of the adults, with more alkanes and less alkenes than males or females related with the artificial diet. As conclusion, the effect of larval substrate on the survival and fecundity parameters is evident. The implications for the mass production of the house fly as animal feed are also indicated.

Low transfer of cadmium, lead and aflatoxin B1 to eggs and meat of laying hens receiving diets with black soldier fly larvae reared on contaminated substrates

Replacing soybeans with insects in egg and poultry meat production could improve environmental sustainability. Black soldier fly larvae (BSFL) have a favorable nutrient composition and can be reared on low-grade waste, but this is associated with the risk of feed and food contamination. The aim of this study was to assess the transfer of selected contaminants from larval substrates to poultry-derived food. Two different control substrates were used. Substrate CCH (produced in Switzerland) was based on side streams approved for insect rearing in the European Union (EU), while substrate CIND (produced in Indonesia) included non-EU approved waste. In addition, substrate CIND was spiked with either heavy metals (HM; 1.9 mg cadmium and 18.8 mg lead/kg dry matter (DM)) or 1.5 mg aflatoxin B1/kg DM (AF)). The larvae fed HM contained 7 mg cadmium and 16 mg lead/kg DM. These values were about 30 times the concentrations of cadmium and 30–60 times the concentrations of lead found on average in the BSFL reared with the two non-spiked substrates. Although substrate AF contained 842 μg aflatoxin B1/kg DM as analysed, the AF larvae contained only 4 μg aflatoxin B1/kg DM. Larval meals were integrated at 200 g/kg in two control diets (diets CCH and CIND) and two diets based on contaminated BSFL (diets HM and AF) designed for late-laying hens (n = 9/treatment). After feeding these diets for 4 weeks, the hens were slaughtered. Diet HM and AF did not affect laying performance or egg quality compared with the control diets. In the body tissue, the cadmium concentrations (per kg DM) were nearly doubled by diet HM in the breast meat (13.3 μg), kidneys (12.3 mg) and liver (1.86 mg) compared to diet CIND. The same diet increased lead in kidneys from below 0.1 to 0.5 mg/kg DM. No lead was detected in the meat and eggs, and no cadmium was found in the eggs. In conclusion, despite cadmium and lead also occurring in BSFL meals of CCH and CIND, the levels in all corresponding hen-based feed and food materials were below the maximum content, except for the kidneys. The aflatoxin B1 level of diet AF (1 μg/kg DM) suggests that the risk might also be small when BSFL are reared on moldy substrate containing aflatoxin-producing fungi. In conclusion, postconsumer waste apparently poses a lower risk than expected in poultry food chains for these contaminants when used as larval substrate.

Yeast and fruit fly mutual niche construction and antagonism against mould

Abstract A goal in insect–microbe ecology is to understand the mechanisms regulating species associations and mutualistic interactions. The spotted wing drosophila Drosophila suzukii develops in ripening fruit, unlike other drosophilids that typically feed on overripe fruit, and is associated with the yeast Hanseniaspora uvarum. We hypothesized that D. suzukii and H. uvarum engage in niche construction leading to a mutualistic relation, facilitating the exploitation of fruit and berries as larval substrate. We show that H. uvarum proliferates on both ripe and on unripe raspberries, mediates attraction of D. suzukii larvae and adult flies, enhances egg‐laying in mated females and is a sufficient food substrate to support larval development. Moreover, H. uvarum suppresses the antagonistic grey mould, Botrytis cinerea in collaboration with D. suzukii larvae, and produces less ethanol than baker's yeast. H. uvarum thus creates favourable conditions for D. suzukii larval development, which is susceptible to ethanol and grey mould. D. suzukii, on the other hand, vectors H. uvarum to suitable substrates such as raspberries, where larval feeding activity enhances growth of H. uvarum. Larval feeding also helps to suppress B. cinerea, which otherwise outcompetes H. uvarum on raspberry, in the absence of fly larvae. In conclusion, H. uvarum enhances D. suzukii larval development on unripe berries, and D. suzukii promotes H. uvarum dispersal and growth on berries. Yeast and fly modify their shared habitat in reciprocal niche construction and mutual interaction. Read the free Plain Language Summary for this article on the Journal blog.

Larval Conditioning and Aging of Sand Fly Rearing Medium Affect Oviposition Site Selection in Phlebotomus papatasi (Diptera: Psychodidae) Sand Flies.

Sand fly larvae develop in sheltered humid habitats containing decaying organic matter on which they feed. Previously, we showed that gravid females of Phlebotomus papatasi Scopoli (Diptera: Psychodidae) are attracted to and stimulated to lay eggs on larval rearing medium containing larvae. That study, however, did not control for the possible effect of medium aging. Our goal in this study was to evaluate the effect of larval substrate conditioning on attraction and oviposition responses of Ph. papatasi sand flies while controlling for the effect of substrate aging. Initially, we confirmed that the pretreatment fresh larval food sources (to be used as larval conditioned and unconditioned media) did not differ with respect to their effect on attraction and oviposition responses. The larval conditioned medium was produced by rearing larvae to the second/third-instar stage over 3 wk using the same larval food source. To produce larval unconditioned medium, the same amount of fresh larval food was added to a control rearing cup that did not contain larvae but was aged under identical time and conditions. Two-choice bioassays were conducted to evaluate gravid female's attraction and oviposition response to larval conditioned and unconditioned media. We found that gravid females were significantly attracted (P < 0.05) to larval conditioned medium when compared with unconditioned medium under the same amount of time and conditions. However, no such difference was found with respect to oviposition response. Both attraction and oviposition responses were significantly increased for larval conditioned and unconditioned media in comparison to the initial fresh larval food source.

Abundance, distribution and substrate association of Hong Kong stag beetles (Coleoptera: Lucanidae) in secondary forests

Abstract Ecological studies conducted on lucanids in Asia are limited, resulting in considerable knowledge gaps in the fundamental ecology of these charismatic species (e.g. larval substrate associations), despite their potential importance in wood decomposition. We conducted 25 transect surveys in secondary forest sites in Hong Kong and sampled wood parameters and site‐level wood availability data to test whether these habitat features significantly affected abundance or occurrence of lucanids. Out of 412 wood fragments sampled, we found two species: Prosopocoilus oweni was broadly distributed across sites (but still uncommon at the level of wood fragments occurring in 6.07%) while Nigidius sinicus was comparatively rarer (occurring in 0.73% of examined wood fragments). The dominant species P. oweni was more likely to appear in wood fragments in later decay stages and can be classified as a secondary forest generalist with respect to wood resource utilisation. The surveys failed to detect four additional lucanid species known in Hong Kong (Neolucanus sinicus, E. gracilis, P. biplagiatus, and Aegus chelifer) potentially because of their rarity or usage of other habitats or microhabitats. Protection of existing secondary forests in Hong Kong is recommended to secure supply of suitable habitat and resources. Results obtained by standardised surveys can furthermore serve as a baseline for future assessment of lucanid population change. This study provides ecological insights into lucanids within secondary forest ecosystems of subtropical and tropical Asia.

The effect of Rhodococcus rhodochrous supplementation on black soldier fly (Diptera: Stratiomyidae) development, nutrition, and waste conversion

Black soldier fly larvae are mass produced globally for use as livestock, poultry, and aquaculture feed. Efforts are continuously seeking processes optimising larval rate of growth, size, and waste conversion as a means to lower cost and increase output. Manipulating microbes in the larval substrate (i.e. fermentation or supplementation) has been demonstrated as a potential solution. However, identifying appropriate microbes for use in this process has been limited. The objective of this study was to determine whether supplementing black soldier fly larvae with the oleaginous microbe Rhodococcus rhodochrous would result in accelerated larval development, increased final larval body size, and increased conversion efficiency. Larvae fed a sterile, or non-sterile, diet treated with R. rhodochrous grew 3× faster than the control by the third day and were approximately 2× larger than the control by the conclusion of the experiment. Conversion rate was 2× greater for the treatments indicating less feed would be needed to achieve maximum weight gain. Protein composition of resulting larvae fed diet supplemented with R. rhodochrous, was significantly different than the control not receiving the microbial infusion. Larvae provided the microbe had 4.20 and 2.79% greater fatty acid composition and short-chained fatty acids, respectively, but lower monounsaturated fatty acids (1.60%) and polyunsaturated fatty acids (2.4%). Furthermore, larvae provided R. rhodochrous produced significantly more proteins related to energy production and storage, as well as muscle development and contraction, while those sans microbe, produced proteins related to stress responses (e.g. heat shock proteins). While, this study yielded positive results for the inclusion of R. rhodochrous as part of the black soldier fly larval diet, additional research is needed to optimise the dose at an industrial scale.

An alternative protocol for rearing of stable fly, Stomoxys calcitrans (Diptera: Muscidae) under laboratory conditions

Stomoxys calcitrans is widely adopted in experiments due to their relevance in animal health. Nevertheless, the rearing of S. calcitrans under laboratory conditions is still a challenge. In this manuscript, we show a new method for rearing the immature stages of S. calcitrans using plant material as only substrate source and sterilized sand for pupae to mimic their natural environment in order to maintain the colonies in the laboratory and for other biological tests. For this purpose, 65 eggs were collected from the colony established in the laboratory and placed into Petri dishes (80 × 15 mm) containing 50 grams of a mixture of alfalfa hay (100 g) and distilled water (200 ml), in a controlled environment (27 ± 1 °C and 70 ± 5 % UR), and the experiment was repeated six times. The mean number of days for egg hatching was 1.4 (± 0.5), the larval period from L1 to L3 was 11.33 days (± 1.75) and the pupal period was 5.83 days (± 0.51). The complete cycle from egg to adult was 18.66 days (± 2.06). The mean larval survival rate, pupation rate, and emergence rate (pupa to adult) were 91.03 % (± 3.14), 88.71 % (± 3.17), and 91.13 % (± 3.0), respectively. The results obtained in the present work indicated that the use of a larval substrate with alfalfa hay allowed the development of immature stages of S. calcitrans in laboratory, which has been demonstrated by the good percentages of larval development, pupation, and adult emergence.

Warm temperature alters the chemical cue preference of Acropora tenuis and Heliopora coerulea larvae

Larvae released into the water column rely on chemical cues from the benthos for successful settlement. However, larval preference for substrates may be affected by rising seawater temperature brought about by global climate change. In this study, we examined the effect of elevated temperature on chemical cue preference by larvae of the scleractinian coral, Acropora tenuis, and the octocoral, Heliopora coerulea, collected from northwestern Philippines. At ambient temperature (28 °C), both H. coerulea and A. tenuis larvae showed preference for substrates containing either crustose coralline algae or crude ethanolic extracts from conspecific or congeneric corals. In contrast, at higher temperature (30 °C), greater preference was shown for substrates containing the crude extract from conspecific or congeneric corals. These results demonstrate that elevated temperature can change larval substrate preference, which will have downstream impacts on crucial biological processes, such as larval settlement and recruitment.

The role of larval substrate specialization and female oviposition in mediating species diversity of closely-related sepsid flies (Diptera: Sepsidae)

Coprophagous insect communities play a critical role in the decomposition of vertebrate dung and provide ecosystem functions fundamental to modern agriculture. While the ecology of dung beetles is rather well understood, niche differentiation in coprophagous flies is poorly studied. Sepsid flies (Diptera: Sepsidae) are a vital part of the European community of coprophages, with 6-7 widespread species of Sepsis often found co-occurring in the same pasture. To advance our ecological understanding of the mechanisms that enable species to coexist, we investigated the oviposition preferences and larval performance of 7 common species of Sepsis in the dung of different large domestic and wild mammals. Substrate preferences and subsequent performance of larvae in laboratory experiments did not vary greatly. All species did very well on cow dung, the most common substrate in Central Europe, but also on dung of horse and wild boar. In contrast, flies did not prefer or grow well in dung of red and roe deer, two of the most common wild vertebrates. Thus there were only minor differences among the species tested along the specialist-generalist (dung) gradient, indicating that differences in the choice of oviposition sites by the adults of the different fly species and larval performance do not constitute a major axis of ecological differentiation. Nevertheless, there was a positive correlation between substrate choice and larval performance indicating the preference of gravid females for particular oviposition sites is adaptive. We conclude that sepsids are common in Europe because they are well adapted to the dung of herbivorous livestock rather than wild animals. Nevertheless, specialization on particular types of dung does not define the niche of Sepsis dung flies and hence plays a minor role in mediating their species diversity.

Insights into the ecology, genetics and distribution of Lucanus elaphus Fabricius (Coleoptera: Lucanidae), North America's giant stag beetle

Abstract Little is known about the biology or conservation status of Lucanus elaphus Fabricius in North America despite well‐documented declines of a related species, Lucanus cervus (L.), in Europe. This study provides information critical to developing conservation plans for L. elaphus including the species’ larval substrate requirements, genetic data and range‐wide estimates of habitat suitability. In Mississippi floodplain forests, larval L. elaphus were recovered from a wide range of log sizes and rot types and were either found tunnelling within the wood or feeding beneath logs at the soil–wood interface. The species appears to require 1–2 years to complete development, exhibits a 1:1 sex ratio and is parasitised by Zelia vertebrata (Say) (Diptera: Tachinidae). Flight intercept traps placed at three heights at both the edge and interior of hardwood‐dominated forests in Georgia yielded six adult male L. elaphus, all of which were captured in traps placed at 15 m on the forest edge. Because L. elaphus larvae are morphologically indistinguishable from related species, DNA sequence data from the mitochondrial cytochrome oxidase I gene were generated to facilitate molecular identification. Genetic data revealed modest intra‐specific variation, with up to 1.3% sequence divergence among haplotypes sampled from the same forest. Based on assembled occurrence records, ecological niche models suggest that environmental conditions are suitable for L. elaphus across much of the southeastern United States, provided that adequate lowland forest cover and dead wood substrates are available.

The influence of substrate material on ascidian larval settlement

Submerged man-made structures present novel habitat for marine organisms and often host communities that differ from those on natural substrates. Although many factors are known to contribute to these differences, few studies have directly examined the influence of substrate material on organism settlement. We quantified larval substrate preferences of two species of ascidians, Ciona intestinalis (cryptogenic, formerly C. intestinalis type B) and Botrylloides violaceus (non-native), on commonly occurring natural (granite) and man-made (concrete, high-density polyethylene, PVC) marine materials in laboratory trials. Larvae exhibited species-specific settlement preferences, but generally settled more often than expected by chance on concrete and HDPE. Variation in settlement between materials may reflect preferences for rougher substrates, or may result from the influence of leached chemicals on ascidian settlement. These findings indicate that an experimental plate material can influence larval behavior and may help us understand how substrate features may contribute to differences in settlement in the field.

Bacterial Associations Across House Fly Life History: Evidence for Transstadial Carriage From Managed Manure.

House flies (Diptera: Muscidae; Musca domestica L.) associate with microbe-rich substrates throughout life history. Because larvae utilize bacteria as a food source, most taxa present in the larval substrate, e.g., manure, are digested or degraded. However, some species survive and are present as third-instar larvae begin pupation. During metamorphosis, many bacteria are again lost during histolysis of the larval gut and subsequent remodeling to produce the gut of the imago. It has been previously demonstrated that some bacterial species survive metamorphosis, being left behind in the puparium, present on the body surface, or in the gut of the emerged adult. We used a combined culture-molecular approach to identify viable microbes from managed manure residue and a wild population of house fly larvae, pupae, puparia, and adults to assess transstadial carriage. All larval (10/10), pupal (10/10), and puparial (10/10) cultures were positive for bacteria. Several bacterial species that were present in larvae also were present either in pupae or puparia. Four viable bacterial species were detectable in 6 of 10 imagoes reared from manure. Of note is the apparent transstadial carriage of Bacillus sonorensis, which has been associated with milk spoilage at dairies, and Alcaligenes faecalis, which can harbor numerous antibiotic resistance genes on farms. The potential of newly emerged flies to harbor and disseminate bacteria from managed manure on farms is an understudied risk that deserves further evaluation.

Niches of Saproxylic Weevils (Coleoptera: Curculionidae) in French Guiana

The loss of tropical forests has a direct effect on saproxylic beetles through the loss of larval feeding substrates and an indirect effect through microclimate change in forest fragments. Impacts will be exacerbated by climate change, and species with narrow niches will be particularly vulnerable. This study presents baseline data on niches (seasonality, host plant, stratum, and branch diameter) of saproxylic weevils in a minimally disturbed moist forest of French Guiana. Weevils (excluding bark beetles) were expected to be host specialists and more abundant in moist microhabitats. Bait branches from three species in the Brazil nut family (Lecythidaceae) were exposed during the dry and rainy season at ground and canopy stratum. They yielded 1,262 weevils in 24 species; 95% belonged to the subfamily Conoderinae. Weevils emerged in greatest abundance from Eschweilera coriacea (A. P. de Candolle) S. A. Mori and from branches in the coolest, moistest microhabitat: ground stratum during the rainy season. Shifts towards warmer, drier climatic conditions would probably have a negative impact on most saproxylic weevil species currently associated with the Brazil nut family. Although this family is expected to suffer under biotic homogenization, the favored host species is hyperdominant throughout Amazonia; climate permitting, E. coriacea could provide a refuge.

Phylogenetics of the antopocerus-modified tarsus clade of Hawaiian Drosophila: diversification across the Hawaiian Islands.

The Hawaiian Drosophilidae radiation is an ecologically and morphologically diverse clade of almost 700 described species. A phylogenetic approach is key to understanding the evolutionary forces that have given rise to this diverse lineage. Here we infer the phylogeny for the antopocerus, modified tarsus and ciliated tarsus (AMC) clade, a lineage comprising 16% (91 of 687 species) of the described Hawaiian Drosophilidae. To improve on previous analyses we constructed the largest dataset to date for the AMC, including a matrix of 15 genes for 68 species. Results strongly support most of the morphologically defined species groups as monophyletic. We explore the correlation of increased diversity in biogeography, sexual selection and ecology on the present day diversity seen in this lineage using a combination of dating methods, rearing records, and distributional data. Molecular dating analyses indicate that AMC lineage started diversifying about 4.4 million years ago, culminating in the present day AMC diversity. We do not find evidence that ecological speciation or sexual selection played a part in generating this diversity, but given the limited number of described larval substrates and secondary sexual characters analyzed we can not rule these factors out entirely. An increased rate of diversification in the AMC is found to overlap with the emergence of multiple islands in the current chain of high islands, specifically Oahu and Kauai.

Effects of terrigenous sediment on settlement and survival of the reef coral Pocillopora damicornis.

Survival and settlement of Pocillopora damicornis larvae on hard surfaces covered with fine (<63 µm) terrigenous red clay was measured in laboratory Petri dishes. The dishes were prepared with sediment films of various thicknesses covering the bottoms. Coral larvae were incubated in the dishes for two weeks and the percent that settled on the bottom was determined. There was a statistically significant relationship between the amount of sediment and coral recruitment on the bottom, with no recruitment on surfaces having a sediment cover above 0.9 mg cm−2. Experimental conditions for the delicate coral larvae were favorable in these experiments. Total survival over the two week settlement tests expressed as the sum of coral recruits and live larvae at the end of the experiment did not show a significant decline, so the major impact of the sediment was on successful settlement rather than on mortality. Larval substrate selection behavior was the primary factor in the observed result.

Biophysical Constraints on Optimal Patch Lengths for Settlement of a Reef-Building Bivalve

Reef-building species form discrete patches atop soft sediments, and reef restoration often involves depositing solid material as a substrate for larval settlement and growth. There have been few theoretical efforts to optimize the physical characteristics of a restored reef patch to achieve high recruitment rates. The delivery of competent larvae to a reef patch is influenced by larval behavior and by physical habitat characteristics such as substrate roughness, patch length, current speed, and water depth. We used a spatial model, the “hitting-distance” model, to identify habitat characteristics that will jointly maximize both the settlement probability and the density of recruits on an oyster reef (Crassostrea virginica). Modeled larval behaviors were based on laboratory observations and included turbulence-induced diving, turbulence-induced passive sinking, and neutral buoyancy. Profiles of currents and turbulence were based on velocity profiles measured in coastal Virginia over four different substrates: natural oyster reefs, mud, and deposited oyster and whelk shell. Settlement probabilities were higher on larger patches, whereas average settler densities were higher on smaller patches. Larvae settled most successfully and had the smallest optimal patch length when diving over rough substrates in shallow water. Water depth was the greatest source of variability, followed by larval behavior, substrate roughness, and tidal current speed. This result suggests that the best way to maximize settlement on restored reefs is to construct patches of optimal length for the water depth, whereas substrate type is less important than expected. Although physical patch characteristics are easy to measure, uncertainty about larval behavior remains an obstacle for predicting settlement patterns. The mechanistic approach presented here could be combined with a spatially explicit metapopulation model to optimize the arrangement of reef patches in an estuary or region for greater sustainability of restored habitats.

Improvement of survival of the house fly (Musca domestica L.) larvae under mass-rearing conditions

Two new approaches were examined, aimed at increasing survival of the house fly (Musca domestica L.) larvae under mass-rearing conditions of a biodegradation facility: modification of the larval substrate and dispersal of the eggs during inoculation. The two types of pig manure used in this study (manure with sawdust and manure without sawdust) differed in terms of larval survival and nutritional value for the house fly larvae. Larval survival in manure without sawdust in the control treatment was low (46.8±2.1%) and its nutritional value for the larvae were high. Addition of 5.7% of previously biodegraded manure did not significantly affect larval survival (52.3±1.9%), but larval development was faster and the pupae were significantly smaller (14.28±0.4mg) compared to the control (16.29±0.5mg). Using alternative substrate for incubation of eggs and first-instar larvae significantly increased larval survival (63.3±3.3%) and decreased the mean weight of produced pupae (14.39±0.71mg). Overall, the weight of recovered biomass in the alternative substrate treatment increased by 14.3kg ton-1 of manure compared to the control. Larval survival in manure with sawdust was generally higher than 70%, but its nutritional value for the larvae was lower than in manure without sawdust. Dispersal of eggs over the surface of manure with sawdust significantly affected larval survival and mean weight of pupae. Larval survival was significantly lower (59.2±4.0%) and pupae were significantly heavier (18.45±0.8mg) when eggs were applied to a small area on the manure surface (spot treatment), as compared to diagonal, Z-line and multiple zig-zag dispersal (72.5±2.4 to 74.6±3.0% and 14.76±0.6 to 15.97±0.6mg, respectively). No significant differences were observed in larval survival or mean weight of pupae when comparing the diagonal, Z-line and multiple zig-zag dispersal patterns. Implementation of the techniques which improve larval survival and increase the weight of produced fly biomass may decrease demand for production of house fly eggs and, therefore, reduce the maintenance costs of adult colony, as well as increase the revenue earned by selling the products.

Decomposed Flesh as a Vitellogenic Protein Source for the Forensically Important &lt;I&gt;Lucilia sericata&lt;/I&gt; (Diptera: Calliphoridae)

For the carrion-feeding blow flies, a common source of vitellogenic protein are the carcasses that also serve as larval substrates. This attraction continues well after the blow flies have ceased to oviposit on the carrion, and it has been assumed that a decomposed cadaver remains a suitable source for vitellogenic protein; however, this assumption has never been tested. To test this hypothesis, blow flies (Lucilia sericata Meigen) were supplied with either fresh or decomposed beef liver as protein sources for vitellogenesis for 5 d. Both treatments produced identical ovarian development, indicating that decomposed flesh remains a suitable protein source for carrion-feeding blow flies. These results support the theory that virgin flies are attracted to cadavers of advanced decomposition as a source of protein.