Jeffery K. Tomberlin

Microbial Volatile Emissions as Insect Semiochemicals

We provide a synthesis of the literature describing biochemical interactions between microorganisms and insects by way of microbial volatile organic compound (MVOC) production. We evaluated the functionality and ecological context of MVOC signals, and explored important metabolic pathways involved in MVOC production. The cosmopolitan distribution of microorganisms creates a context for frequent, and frequently overlooked, insect responses to microbial emissions. There are numerous instances of MVOCs being closely associated with insect feeding behaviors, but some MVOCs are also powerful repellants. Emissions from microorganisms in situ may signal aspects of habitat suitability or potential exposure to entomopathogens. In some ecosystems, bacterial or fungal volatiles can also incite insect aggregations, or MVOCs can resemble sexual pheromones that elicit mating and oviposition behaviors from responding insects. A single microorganism or MVOC can have different effects on insect behaviors, especially across species, ontogenies, and habitats. There appears to be a multipartite basis for insect responses to MVOCs, and complex tritrophic interactions can result from the production of MVOCs. Many biochemical pathways for behaviorally active volatile production by microbial species are conserved across large taxonomic groupings of microorganisms. In addition, there is substantial functional redundancy in MVOCs: fungal tissues commonly produce polyketides and short-chain alcohols, whereas bacterial tissues tend to be more commonly associated with amines and pyrazines. We hypothesize that insect olfactory responses to emissions from microorganisms inhabiting their sensory environment are much more common than currently recognized, and that these signals represent evolutionarily reliable infochemicals. Insect chemoreception of microbial volatiles may contribute to the formation of neutral, beneficial, or even harmful symbioses and provide considerable insight into the evolution of insect behavioral responses to volatile compounds.

A Roadmap for Bridging Basic and Applied Research in Forensic Entomology

The National Research Council issued a report in 2009 that heavily criticized the forensic sciences. The report made several recommendations that if addressed would allow the forensic sciences to develop a stronger scientific foundation. We suggest a roadmap for decomposition ecology and forensic entomology hinging on a framework built on basic research concepts in ecology, evolution, and genetics. Unifying both basic and applied research fields under a common umbrella of terminology and structure would facilitate communication in the field and the production of scientific results. It would also help to identify novel research areas leading to a better understanding of principal underpinnings governing ecosystem structure, function, and evolution while increasing the accuracy of and ability to interpret entomological evidence collected from crime scenes. By following the proposed roadmap, a bridge can be built between basic and applied decomposition ecology research, culminating in science that could withstand the rigors of emerging legal and cultural expectations.

Rearing methods for the black soldier fly (Diptera: Stratiomyidae).

Abstract The black soldier fly, Hermetia illucens (L.), is a nonpest tropical and warm-temperate region insect that is useful for managing large concentrations of animal manure and other biosolids. Manure management relying on wild fly oviposition has been successful in several studies. However, confidence in this robust natural system was low and biological studies were hampered by the lack of a dependable source of eggs and larvae. Larvae had been reared easily by earlier investigators, but achieving mating had been problematic. We achieved mating reliably in a 2 by 2 by 4-m screen cage in a 7 by 9 by 5-m greenhouse where sunlight and adequate space for aerial mating were available. Mating occurred during the shortest days of winter if the sun was not obscured by clouds. Adults were provided with water, but no food was required. Techniques for egg collection and larval rearing are given. Larvae were fed a moist mixture of wheat bran, corn meal, and alfalfa meal. This culture has been maintained for 3 yr. Maintainance of a black soldier fly laboratory colony will allow for development of manure management systems in fully enclosed animal housing and in colder regions.

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

Abstract Hermetia 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 field-collected prepupae and corresponding emergent adults. Diet did not significantly influence development or survivorship to the prepupal stage. However, adult emergence for all diets was significantly less than that determined for the wild population. Development time from egg to adult for individuals reared on the diets at 27°C ranged from 40 to 43 d with the larval stage lasting 22–24 d. We observed >96% larval survivorship to the prepupal stage and 21–27% adult emergence. Females accounted for 55–60% 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 field-collected specimens were significantly smaller than females and emerged 1–2 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 flies. However, further refinement is needed to produce adults similar to those found in nature.

Development of Black Soldier Fly (Diptera: Stratiomyidae) Larvae Fed Dairy Manure

Abstract Black soldier flies, Hermetia illucens L., are a common colonizer of animal wastes. However, all published development data for this species are from studies using artificial diets. This study represents the first examining black soldier fly development on animal wastes. Additionally, this study examined the ability of black soldier fly larvae to reduce dry matter and associated nutrients in manure. Black soldier fly larvae were fed four rates of dairy manure to determine their effects on larval and adult life history traits. Feed rate affected larval and adult development. Those fed less ration daily weighed less than those fed a greater ration. Additionally, larvae provided the least amount of dairy manure took longer to develop to the prepupal stage; however, they needed less time to reach the adult stage. Adults resulting from larvae provided 27 g dairy manure/d lived 3–4 d less than those fed 70 g dairy manure. Percentage survivorship to the prepupal or adult stages did not differ across treatments. Larvae fed 27 g dairy manure daily reduced manure dry matter mass by 58%, whereas those fed 70 g daily reduced dry matter 33%. Black soldier fly larvae were able to reduce available P by 61–70% and N by 30–50% across treatments. Based on results from this study, the black soldier fly could be used to reduce wastes and associated nutrients in confined bovine facilities.

Bioconversion of dairy manure by black soldier fly (Diptera: Stratiomyidae) for biodiesel and sugar production

Modern dairies cause the accumulation of considerable quantity of dairy manure which is a potential hazard to the environment. Dairy manure can also act as a principal larval resource for many insects such as the black soldier fly, Hermetia illucens. The black soldier fly larvae (BSFL) are considered as a new biotechnology to convert dairy manure into biodiesel and sugar. BSFL are a common colonizer of large variety of decomposing organic material in temperate and tropical areas. Adults do not need to be fed, except to take water, and acquired enough nutrition during larval development for reproduction. Dairy manure treated by BSFL is an economical way in animal facilities. Grease could be extracted from BSFL by petroleum ether, and then be treated with a two-step method to produce biodiesel. The digested dairy manure was hydrolyzed into sugar. In this study, approximately 1248.6g fresh dairy manure was converted into 273.4 g dry residue by 1200 BSFL in 21 days. Approximately 15.8 g of biodiesel was gained from 70.8 g dry BSFL, and 96.2g sugar was obtained from the digested dairy manure. The residual dry BSFL after grease extraction can be used as protein feedstuff.

Development of the Black Soldier Fly (Diptera: Stratiomyidae) in Relation to Temperature

ABSTRACT The black soldier fly, Hermetia illucens L., was reared on a grain-based diet at 27, 30, and 36°C. Survival of 4- to 6-d-old larvae to adults averaged 74–97% at 27 and 30°C but was only 0.1% at 36°C. Flies required a mean of ≈4 d (11%) longer to complete larval and pupal development at 27°C than at 30°C. At 27 and 30°C, females weighed an average of 17–19% more than males but required an average of 0.6–0.8 d (3.0–4.3%) longer to complete larval development. At both temperatures, adult females lived an average of ≈3.5 d less than adult males. The duration of larval development was a significant predictor of adult longevity. Temperature differences of even 3°C produce significant fitness tradeoffs for males and females, influencing life history attributes and having practical applications for forensic entomology.

Proteus mirabilis interkingdom swarming signals attract blow flies

Flies transport specific bacteria with their larvae that provide a wider range of nutrients for those bacteria. Our hypothesis was that this symbiotic interaction may depend on interkingdom signaling. We obtained Proteus mirabilis from the salivary glands of the blow fly Lucilia sericata; this strain swarmed significantly and produced a strong odor that attracts blow flies. To identify the putative interkingdom signals for the bacterium and flies, we reasoned that as swarming is used by this bacterium to cover the food resource and requires bacterial signaling, the same bacterial signals used for swarming may be used to communicate with blow flies. Using transposon mutagenesis, we identified six novel genes for swarming (ureR, fis, hybG, zapB, fadE and PROSTU_03490), then, confirming our hypothesis, we discovered that fly attractants, lactic acid, phenol, NaOH, KOH and ammonia, restore swarming for cells with the swarming mutations. Hence, compounds produced by the bacterium that attract flies also are utilized for swarming. In addition, bacteria with the swarming mutation rfaL attracted fewer blow flies and reduced the number of eggs laid by the flies. Therefore, we have identified several interkingdom signals between P. mirabilis and blow flies.

Black soldier fly (Diptera: Stratiomyidae) larvae reduce Escherichia coli in dairy manure.

Abstract Escherichia coli labeled with a green fluorescent protein was inoculated into sterile dairy manure at 7.0 log cfu/g. Approximately 125 black soldier fly larvae were placed in manure inoculated and homogenized with E. coli. Manure inoculated with E. coli but without black soldier fly larvae served as the control. For the first experiment, larvae were introduced into 50, 75, 100, or 125 g sterilized dairy manure inoculated and homogenized with E. coli and stored 72 h at 27°C. Black soldier fly larvae significantly reduced E. coli counts in all treatments. However, varying the amount of manure provided the black soldier fly larvae significantly affected their weight gain and their ability to reduce E. coli populations present. For the second experiment, larvae were introduced into 50 g manure inoculated with E. coli and stored for 72 h at 23, 27, 31, or 35°C. Minimal bacterial growth was recorded in the control held at 35°C and was excluded from the analysis. Black soldier fly larvae significantly reduced E. coli counts in manure held at remaining temperatures. Accordingly, temperature significantly influenced the ability of black soldier fly larvae to develop and reduce E. coli counts with greatest suppression occurring at 27°C.

Microbial community functional change during vertebrate carrion decomposition.

Microorganisms play a critical role in the decomposition of organic matter, which contributes to energy and nutrient transformation in every ecosystem. Yet, little is known about the functional activity of epinecrotic microbial communities associated with carrion. The objective of this study was to provide a description of the carrion associated microbial community functional activity using differential carbon source use throughout decomposition over seasons, between years and when microbial communities were isolated from eukaryotic colonizers (e.g., necrophagous insects). Additionally, microbial communities were identified at the phyletic level using high throughput sequencing during a single study. We hypothesized that carrion microbial community functional profiles would change over the duration of decomposition, and that this change would depend on season, year and presence of necrophagous insect colonization. Biolog EcoPlates™ were used to measure the variation in epinecrotic microbial community function by the differential use of 29 carbon sources throughout vertebrate carrion decomposition. Pyrosequencing was used to describe the bacterial community composition in one experiment to identify key phyla associated with community functional changes. Overall, microbial functional activity increased throughout decomposition in spring, summer and winter while it decreased in autumn. Additionally, microbial functional activity was higher in 2011 when necrophagous arthropod colonizer effects were tested. There were inconsistent trends in the microbial function of communities isolated from remains colonized by necrophagous insects between 2010 and 2011, suggesting a greater need for a mechanistic understanding of the process. These data indicate that functional analyses can be implemented in carrion studies and will be important in understanding the influence of microbial communities on an essential ecosystem process, carrion decomposition.