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United States Department of Agriculture

Agricultural Research Service

Research Project: Mass Production of Biological Control Agents

Location: Biological Control of Pests Research Unit

2012 Annual Report


1a.Objectives (from AD-416):
The long-term objective of this project is to develop an improved understanding of the biology, nutrition, and behavior of natural enemies of pest arthropods. Our primary target pest will be the two-spotted spider mite, Tetranychus urticae. Some of the most important natural enemies of this mite are phytoseiid predatory mites and coccinellids. We aim to implement, improve and increase the mass rearing capabilities of these organisms by developing artificial diets and novel rearing technology to enable economical biological control of this pest by augmentation of its natural enemies and to develop novel techniques of quality control and quality assurance. Our work will focus on the following: (1) Determine nutritional, behavioral, and physical conditions necessary for in vivo and in vitro insect rearing and use the results to implement mass production systems of biological control agents. (2) Determine the impact of mass rearing procedures on survival, reproduction, and gene expression. (3) Assess optimal conditioning of insects for release.


1b.Approach (from AD-416):
Research activities will focus on a specific predator, Stethorus punctillum and a generalistic predator, Coleomegilla (C.) maculata. Studying the nutritional requirements of a generalist versus a specific predator will provide insights on diet development of these 2 organisms. The generalist predator is expected to be an easier target for artificial diet development and success will have greater impact since C. maculate is an important predator of several economically damaging pest species.


3.Progress Report:
Three artificial diet formulations have been developed for the pink spotted lady beetle, Coleomegilla (C.) maculata. Full diet evaluation and comparison to a control consisting of bee pollen and Ephestia kuehniella eggs (1:1) is underway and expected to be completed before the end of the fiscal year. Lady beetles have completed development and reproduced in each of the 3 diet formulations. Three new cage designs have been introduced to improve C. maculata rearing. One of the designs improves egg collection and reduces egg cannibalism; the second design allows immature rearing in large groups using artificial diet, reducing cannibalism; in the third design, large numbers of eggs are collected from hundreds of adults fed with artificial diet where oviposition and feeding areas are divided to prevent cannibalism. Temperature dependent development studies of C. maculata at 3ºC intervals are under way and expected to be completed before the end of September. Studies on the life history of the pink spotted lady beetle revealed that this predator readily consumes all life stages of the two-spotted spider mite (Tetranychus (T.) urticae) in rearing systems under development. Plans are underway to test the foraging rate of predators (searching for spider mites) on live plants in rearing rooms and a greenhouse. A highly inbred colony with wild type phenotype was selected from the stock colony and a yellow color morph was obtained by laboratory inbreeding. A colony is currently under selection for a lighter red/gold phenotype. Olfactometer studies demonstrated that the plant-derived molecule methyl salicylate was attractive to C. maculata adults but did not stimulate egg production or feeding. Ongoing research is examining other molecules for their potential as oviposition and/or feeding stimulants. Experiments are in progress to define the influence of extreme temperature and humidity that predators (C. maculata) endure while transported from commercial producers to end-users (farmers, growers). Preliminary observations done with mini loggers indicate that temperatures fluctuate widely during transport. These fluctuations have negative consequences on the fecundity (reproduction) of predators at the point of release for augmentative biological control. In the in-vivo rearing of nematodes, diets containing 0.15% manganese and 0.16% cholesterol fed to yellow mealworms improved virulence in one nematode species. A multiple-choice study using 7 diet ingredients resulted in a better diet for the yellow mealworm using self-selection to adjust ingredient ratios. Successful production of the spined soldier bug (Podisus maculiventris) eggs allowed continuous culture of 4 imported and 4 native egg parasitoid species being evaluated for application against 3 exotic invasive insects: The brown marmorated stink bug, the bagrada bug and the kudzu bug.


4.Accomplishments
1. Artificial diets for Coleomegilla (C.) maculata. Current rearing techniques of the twelve spotted lady beetle C. maculata consist of using mixtures of pollen and eggs from the Mediterranean flour moth (Ephestia kuehniella). This method is expensive and labor intensive. ARS scientists in Biological Control of Pests Research Unit at Stoneville, MS, developed 3 artificial diet formulations that allow the twelve spotted lady beetle to complete development and reproduce. Colonies have been maintained solely on artificial diet for more than 10 generations without the loss of vigor. These new formulations can reduce the costs of mass producing these lady beetles allowing their commercial application in the control of several pests including aphids, scales, spider mites, Colorado potato beetles, and more.

2. Nematode virulence enhancing host diets. Commercial production of insect killing (entomopathogenic) nematodes is mostly done by highly sophisticated and expensive fermentors. Nematodes produced by this method often lose virulence and must be replaced. Producing nematodes in-vivo eliminates this problem and also allows production of other nematode species that do not grow in fermentors. Using the yellow mealworm as host for the nematodes is commercially viable, but this beetle produces fewer numbers of nematodes per larva than other species more difficult to produce such as wax worms. ARS scientists in Biological Control of Pests Research Unit at Stoneville, MS, in cooperation with scientists at Byron, GA, improved nematode virulence by modifying the mealworm diet with the addition of manganese sulfate and cholesterol. This diet allows for more nematodes to be produced per mealworm larvae, increasing production and improving the product.

3. Pink spotted lady beetle as a predator of the two spotted spider mite. The pink spotted lady beetle (Colemegilla (C.) maculata) has been reported to feed on the two spotted spider mite, but the potential of this predator to control spider mites has not been demonstrated. The two spotted spider mite is one of the most important pests in greenhouses and it is difficult to control with pesticides due to its wide-range acaricide resistance. ARS scientists in the Biological Control of Pests Research Unit at Stoneville, MS, discovered that the two-spotted spider mite (Tetranychus (T.) urticae) is a highly palatable prey of C. maculata. However, continuous rearing of predators throughout their life cycle on this spider mite has negative consequences on predator growth and reproduction. Predators require a longer time to mature, are smaller – sized adults, and females produce fewer progeny when reared on spider mites rather than on moth eggs. Although C. maculata does not do well when reared on spider mites, this predator potentially can still be used effectively to control spider mite infestations when released in greenhouse crops. This could provide of a new target for this biological control agent and improve its commercial value.

4. Pigmentation deficient strain “Yellow” of pink spotted lady beetle. Colors and patterning of lady beetles have been studied for decades, primarily using the invasive Asian multicolored lady beetle. There have been no studies describing the pigmentation in the twelve spotted lady beetle, nor molecular genetic studies of the biochemical synthesis and translocation of lady beetle pigmentation. ARS scientist in the Biological Control of Pests Research Unit at Stoneville, MS, produced a highly inbred strain of twelve spotted lady beetle, Coleomegilla maculata for genetic sequencing and experimentation that is homozygous for a recessive allele that results in lack of red pigmentation in the elytra and eyes. This insect colony may be used as a genetic marker strain for this environmentally and agriculturally important insect. Further it may be used to study pigment interactions with the environment, particularly pertaining to climate change, and the biochemistry of pigment synthesis with regard to vision and warning coloration in a generalist predator.

5. Rearing the kudzu bug and their egg parasitoids from Japan. The kudzu bug, Megacopta (M.) cribraria, recently became established in Georgia and is now in 7 southeastern states, already damaging soybeans. A classical biocontrol project was begun in the Stoneville Research Quarantine Facility, Stoneville, MS, with the importation of 2 egg parasitoids from Japan. The kudzu bug proved very difficult to rear, so regular shipments of adults were received from collaborators with the University of Georgia and Clemson University to produce eggs for parasitoid rearing and non-target studies with native bugs. It was found that M. cribraria eggs could be frozen at minus 80 degrees Celsius and still be attacked by the parasitoids. Host range studies showed that one species of parasitoid could not successfully attack any native bugs tested.


Review Publications
Riddick, E.W., Rojas, M.G., Wu, Z. 2011. Lima bean lady beetle interactions: spider mite mediates sublethal effects of its host plant on growth and development of its predator. Arthropod-Plant Interactions. 5(4):287-296.

Allen, M.L., Walker III, W.B. 2012. Saliva of Lygus lineolaris digests double stranded ribonucleic acids. Journal of Insect Physiology. 58:391-396.

Morales Ramos, J.A., Rojas, M.G., Shapiro Ilan, D.I., Tedders, W.L. 2011. Self-selection of two diet components by Tennebrio molitor (Coleoptera: Tenebrionidae) larvae and its impact on fitness. Environmental Entomology. 40(50): 1285-1294.

Morales Ramos, J.A., Rojas, M.G., Kay, S., Tedders, W., Shapiro Ilan, D.I. 2012. Impact of adult weight, density, and age on reproduction of Tenebrio molitor (Coleoptera: Tenebrionidae). Journal of Entomological Science. 47:208-220.

Riddick, E.W., Wu, Z. 2012. Mother-offspring relations: prey quality and maternal size affect egg size of an acariphagous lady beetle in culture. Psyche. volume 2012, article ID 764350, 7p.

Roy, H., Rhule, E., Harding, S., Lawson-Handley, L., Poland, R., Riddick, E.W., Steenberg, T. 2011. Living with the enemy: parasites and pathogens of the invasive alien ladybird Harmonia axyridis. BioControl. 56(4):663-679.

Sloggett, J.J., Magro, A., Verheggen, F.J., Hemptinne, J., Hutchison, W.D., Riddick, E.W. 2011. The chemical ecology of Harmonia axyridis. Biocontrol. 56(4):643-661.

Last Modified: 4/20/2014
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