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                    U.S. DEPARTMENT OF AGRICULTURE
 Biological Control of Pests Research: Stoneville, MS
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ARS Home Southeast Area Stoneville, Mississippi Biological Control of Pests Research Research





- Morales Ramos, Juan
- Rojas, Maria - Guadalupe
- Riddick, Eric
- 2023
- 2022
- 2021
- Publications
- Sustainable Agricultural Systems Research (216)
- Crop Protection & Quarantine (304)

Research Project: Mass Production of Insects for Biological Control of Arthropod Pests and as Food Sources **Location:
                            Biological Control of Pests Research** 2023 Annual Report **Objectives**
                                

1. Expand studies on discovery, culturing, production and evaluation of natural enemies for biological control and insects as animal feed.
1.a. Behavior and evaluation of black soldier fly as feed and fertilizer.
1.b. Characterization of traits for high efficiency of food conversion and development of Tenebrio molitor for mass production as animal feed.

2. Develop improved rearing methods and artificial diets to reduce costs of insect mass production.
2.a. Test of prey recognition and consumption rate of lady beetles reared on artificial diet or factitious food for multiple generations.
2.b. Determine the effect of diet on the nutritional value of Tenebrio molitor produced for animal feed. **Approach**
                                

The invertebrate predator Coleomegilla maculata De Geer (Coleoptera: Coccinellidae) will be used as a model invertebrate in this test. It is an omnivorous predator that has the capacity to utilize microalgae and synthetic pollen as food to obtain essential nutrients not present in unsuitable prey (Riddick and Wu, 2015). This colony has been in continuous culture without any addition of wild type individuals for more than 50 generations in a climate-controlled room (24°C, 50%–60% RH, 16 h photophase). Individuals originating from the colony and used in this experiment have been mass-reared for more than 10 consecutive generations on a proprietary mixture of freeze-dried, powdered brine shrimp eggs Artemia franciscana Kellogg, freshwater microalgae (Chlorella vulgaris Beijerink), and fatty acids, stearic acid (18:0) or myristic acid (14:0) (Riddick, unpublished data). Larvae and adults have no prior exposure to H. illucens. Yet, published work using larvae and adults from this colony, indicated that the eggs of the common housefly Musca domestica L. (Diptera: Muscidae) were suitable prey in experimental trials (Riddick et al. 2014). All life stages have been maintained in separate cages in the same climate-controlled room (24 °C, 50%–60% RH, 16 h photophase) for more than 10 years at the National Biological Control Laboratory (NBCL), USDA, ARS, in Stoneville, Mississippi.

Cultivated strawberry (Fragaria × ananessa Duch. cv. Chandler) will serve as model plants to test the effects of H. illucens larval frass, as a fertilizer, on plant health. Approximately 1,000 bare root individuals will be purchased from a commercial nursery in Redding, CA, as mentioned in previous research (Riddick et al. 2019). All individuals will be removed from cold storage in NBCL, then transplanted into “flats” over the course of several consecutive days. Excess dead roots will be trimmed away, using stainless steel scissors. Once in flats at a spacing of approximately 6 cm between individual plants, a light covering of peat soilless media will be applied over the roots; then watered using tap water. All plants will be setup on metal benches in a temperature-controlled greenhouse (20-22°C, natural lighting) in Stoneville, Mississippi, USA. One month later, the healthiest plants will be transferred to plastic (1.3 L) pots, one plant per pot, containing the same quantity of soilless media in each pot. **Progress Report**
                                

Molecular and morphological analyses were conducted to determine if a new Aphelinus species was discovered parasitizing the small bramble aphid (Aphis ruborum) on cultivated strawberry in Stoneville, Mississippi. A molecular phylogeny of the genus Aphelinus based on 8,296 universal orthogroups placed Aphelinus material from Mississippi in the A. varipes complex in a clade that includes A. certus and A. nigritus. However, it was sufficiently different from them that it appeared to represent a new species. Males and females from Mississippi were scored for 62 morphological characters used to differentiate species in Aphelinus. With respect to morphology, they were quite different from Aphelinus nigritus, the only species in this complex thought to be originally from North America, and they also differed from populations of Aphelinus certus collected in Korea, China, and Japan. Mating compatibility experiments between the proposed new cryptic species and previously known cryptic species, e.g., A. certus and A. nigritus, will confirm the results of the morphological and molecular analyses.

ARS researchers discovered that adults of the pink spotted lady beetle recognize and readily consume aphid prey after making physical contact with aphids regardless of the rearing diet (factitious diet, artificial diet, or plant pollen). Ongoing research will compare lady beetle recognition of two aphid species, tentatively identified as oleander aphid (Aphis nerii), fleabane aphid (Uroleucon erigeronense) and (Aulacorthum solani). Note that the aphid species (Aphis ruborum) continuously on strawberry for several years, has been maintained at undetectable levels of abundance in three high tunnels, since monthly adult releases of pink spotted lady beetles began. Publication of this research is forthcoming.

Laboratory experiments revealed that Black Soldier Fly (BSF) larvae, in a powdered form, has limited capacity as a protein supplement in a factitious food and an artificial diet for growth and development of predatory pink spotted lady beetle (Coleomegilla maculata). Larvae developed faster and attained larger adult body mass when reared on a factitious food, e.g., brine shrimp eggs, Chlorella algae, and myristic acid, rather than BSF alone, or BSF plus Chlorella and myristic acid. Similarly, larval development, growth, and adult body size was better on the brine shrimp mixture, that than an artificial diet (powdered formulation), or a 50/50 mixture of BSF and the artificial diet. Interestingly, reproductive output, i.e., lady beetle eggs per clutch or total number of clutches, did not differ significantly between BSF, artificial diet, and the brine shrimp mixture. 	

The colony of the pink spotted lady beetle that has been maintained in artificial diet for over 50 generations. Modifications were done to the current diet formulation by replacing the mealworm powder by house fly pupa powder. This substitution was sufficient to impact the colony positively. Black soldier fly was used to substitute for house fly powder in the diet in combination with mealworm pupae powder. Egg hatching and larval survival levels remain at normal levels and adult survival has improved. These findings support the hypothesis that addition of black soldier fly powder to the artificial diet in a gel formulation provides benefits. The black soldier fly is commercially produced, and its cost is considerably lower than house fly reducing the costs of the lady beetle diet.

Work supporting the grand challenge included the production of 50 kg of yellow mealworm frass at NBCL (National Biological Control Laboratory), Stoneville, Mississippi. The frass was given to our collaborators at the USDA-ARS PPPSRU (Poultry Production and Product Safety Research Unit), Fayetteville, Arkansas, to be chemically analyzed and evaluated as plant fertilizer. Filed evaluations have been completed and the manuscript is in preparation. 50 kg, yellow mealworm powder and house cricket each, were also provided to our collaborators at USDA-ARS PPPSRU to be incorporated into broilers feed formulations at different levels. Experiments have been concluded and data is currently being analyzed.

Also, as part of the grand challenge, mealworm larvae from 4 selected lines were grown in the NBCL at Stoneville, Mississippi. These lines were selected for shorter development time and higher pupal size. Specimens from each of the 4 lines were fixed in a solution and shipped to our cooperators at USDA-ARS Center for Grain and Animal Science Research, Manhattan, Kansas, for genetic analysis. This is part of a study for genetic characterization of multiple populations of the yellow mealworm from diverse origins. We expect to obtain clues on the genetic basis of two quantitative traits (development time and pupal size), which are relevant to biomass productivity. **Accomplishments**
                                

1. Selecting mealworm lines for improved biomass productivity. The yellow mealworm is one of the most important industrialized insects in the world and has a great potential to be used as a substitute for fish meal in animal diets as a source of protein and essential nutrients. Fish meal is becoming increasingly unsustainable due to many factors including overfishing and global warming. Mealworm powder could provide a good substitute if its current price could be reduced. One way to reduce costs of production is to increase its productivity. An ARS researcher in Stoneville, Mississippi, determined that two traits associated with biomass productivity (developmental speed and body size) have sufficient genetic basis to respond to selection. In subsequent studies, four lines were selected at 2 levels of developmental speed and two levels of body size for three generations. This study resulted in significant differences in development time and pupal size among the 4 selected lines. This demonstrated the positive and significant impact of selection on improving mealworm productivity providing the basis for developing commercial improved lines. The potential benefits include creation of improved mealworm lines of increased productivity. Commercial producers of mealworms will potentially benefit from these lines. **Review Publications**
                            *Morales Ramos, J.A., Rojas, M.G., Tweedy, D., Kelstrup, H.C., Emery, V. 2022. Inheritance of two traits with high plasticity, developmental speed and body size, in Tenebrio molitor (Coleoptera: Tenebrionidae). Journal of Insect Science. 22(6):13. https://doi.org/10.1093/jisesa/ieac075.*
                                *Rojas, M.G., Morales Ramos, J.A. 2023. Effects of taurine dietary supplement on the biological and demographic parameters of Nezara viridula (Heteroptera: Pentatomidae). Journal of Insect Science. 23(1):6. https://doi.org/10.1093/jisesa/iead003.*
                                *De Clercq, P., Courdron, T.A., Riddick, E.W. 2023. Production of heteropteran predators. In: Morales-Ramos, J.A., Rogas, M.G., Shapiro-Ilan, D.I., editors. Mass Production of Beneficial Organisms. 2nd edition. San Diego, CA: Academic Press. p. 57-100.*
                                *Riddick, E.W., Wu, Z., Chen, J. 2023. Differential susceptibility of Coleomegilla maculata (Coleoptera: Coccinellidae) and Scymnus creperus (Coleoptera: Coccinellidae) larvae to aggression by Solenopsis invicta (Hymenoptera: Formicidae) workers. Insects. https://doi.org/10.3390/insects14040318.*
                                *Leppla, N.C., Morales Ramos, J.A., Shapiro Ilan, D.I., Rojas, M.G. 2022. Introduction. In: Morales-Ramos, J.A., Rojas, M.G., Shapiro-Ilan, D.A., editors. Mass Production of Beneficial Organisms: Invertebrates and Entomopathogens. 2nd edition. San Diego, CA: Academic Press. p.3-12.*
                                *Morales Ramos, J.A., Goolsby, J., Geden, C.J., Rojas, M.G., Garcia-Cancino, M.D., Rodriguez-Velez, B., Arredondo-Bernal, H., Ciomperlik, M.A., Simmons, G.S., Gould, J.R., Hoelmer, K.A. 2022. Production of hymenopteran parasitoids. In: Morales-Ramos, J.A., Rojas, M.G., Shapiro-Ilan, D.I., editors. Mass Production of Beneficial Organisms: Invertebrates and Entomopathogens. 2nd edition. San Diego, CA: Academic Press. p. 101-155.*
                                *Morales Ramos, J.A., Rojas, M.G., Coudron, T.A., Huynh, M.P., Zou, D., Shelby, K. 2022. Artificial diet development for entomophagous arthropods. In: Morales-Ramos, J.A., Rojas, M.G., Shapiro-Ilan, D.I., editors. Mass Production of Beneficial Organisms: Invertebrates and Entomopathogens. 2nd edition. San Diego, CA: Academic Press. p.233-260.*
                                *Riddick, E.W. 2023. Production of Coleopteran predators. In: Morales-Ramos, J.A., Rojas, M.G., Shapiro-Ilan, D.I., editors. Mass Production of Beneficial Organisms: Invertebrates and Entomopathogens. 2nd edition. San Diego, CA: Academic Press. p. 13-36.*


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