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ARS Home » Northeast Area » Ithaca, New York » Robert W. Holley Center for Agriculture & Health » Emerging Pests and Pathogens Research » Research » Research Project #429999

Research Project: Microbial and Arthropod Biological Control Agents for Management of Insect Pests of Greenhouse Crops and Trees

Location: Emerging Pests and Pathogens Research

2018 Annual Report

Objective 1: Integrate mycoinsecticides to manage selected insect pests of greenhouse crops, including, but not limited to, soil and foliar treatments for thrips control and for optimizing abiotic conditions. (NP304, Component 3, Problem Statement 3A2) Subobjectives: 1a. Characterize and quantify the effects of biotic and abiotic factors on efficacy of fungal pathogens applied against western flower thrips. 1b. Assess the potential of combining soil and foliar applications of fungi against western flower thrips as a strategy to achieve acceptable levels of efficacy at reduced moisture levels. Objective 2: Characterize pest microbial associates and determine the efficacy of microbial control agents to manage insect pests, including, but not limited to, Asian ambrosia beetles, walnut twig beetle, and coffee berry borer. Subobjectives: 2a. Determine effects of commercially available fungal biocontrol agents on Asian ambrosia beetles and their symbionts. 2b. Assess efficacy of commercially available entomopathogenic fungi for the walnut twig beetle. 2c. Assess efficacy of Beauveria bassiana against coffee berry borer in the Kona coffee-growing region on the island of Hawaii; determine best practices for effective deployment of this pathogen as one component of an area wide IPM program. 2d. Characterize the indigenous isolates of Beauveria infecting coffee berry borer in Hawaii and determine the roles of these pathogens in natural suppression of this insect pest. Objective 3: Determine the impact of natural enemies, such as parasitic wasps and microbial biopesticides, introduced for management of emerald ash borer. Subobjectives: 3a. Quantify EAB and parasitoid densities to determine whether establishment of EAB parasitoids has significant effects on EAB populations. 3b. Determine impact of natural enemies released against emerald ash borer on ash health and survival.

The goals of this project are to integrate entomopathogenic fungi into management systems for insect pests of greenhouse ornamental and vegetable crops and insect pests of trees in nursery, field, and natural settings and to track fungal strains in these environments. The work comprises fundamental laboratory studies as well as applied field and greenhouse research. This project will develop basic information on the biology of fungal pathogens associated with insects, their genetic and phenotypic variability, and their activity and persistence in field and greenhouse environments. Integration of fungal pathogens will be accomplished for management of key pests. Studies will identify minimal ambient moisture requirements for effective deployment of fungal pathogens against western flower thrips infesting greenhouse crops. Microbial control agents will be developed for management of Asian ambrosia beetles and walnut twig beetle. Application methods and strategies will be developed for optimal use of fungi against coffee berry borer in the Kona coffee districts of Hawaii. Parasitoid releases for management of emerald ash borer will be evaluated for their impact on their host and the resulting impacts on ash health and regeneration. Biological control agents used in the ways developed in this project will provide safe, effective biological alternatives to synthetic chemical insecticides or as rotational partners for insecticide resistance management.

Progress Report
Objective 1a. Laboratory studies to determine the minimum effective moisture regimes for fungal infection of western flower thrips under constant and fluctuating temperature regimes have been completed (see accomplishments). Results indicated that, under constant 25 degrees Celsius or daily fluctuating temperatures of 15–25 degrees Celsius, around 40 hours of high humidity was sufficient to support high levels of infection by biocontrol fungi. Greenhouse pest managers can apply this tactic by making spray applications during late afternoon and maintain high humidity conditions during a period comprising the subsequent night (when high humidity is most easily maintained), the next day (when maintenance of high humidity is most difficult), and the second night. Objective 2a. The Asian ambrosia beetles Xylosandrus germanus (black stem borer), Xylosandrus crassiusculus (granulate ambrosia beetle), and Cnestus mutilatus (camphor shot beetle) are invasive pests that attack a wide range of woody host plants, including many orchard and nursery trees. Research on control of these pests have been conducted by ARS scientists in Ithaca, New York, in collaboration with entomologists from Cornell University and the University of Tennessee. Results from controlled experiments conducted under field conditions have indicated that spray applications of entomopathogenic fungi can significantly impact populations of foundress female beetles and their broods and that mycoparasitic fungi can kill or suppress the associated fungal symbionts these beetles grow in their galleries for food, reducing or eliminating brood production. Testing of fungal pathogens in commercial nursery and orchard crops has been delayed due to a critical SY (Scientist) vacancy; however, collaborative efforts with Cornell scientists are continuing. Insect-repellent products, developed for other systems, are being tested alone and in combination with novel chemical insecticides on stressed potted apple saplings placed in commercial orchards. Potted apple trees, flooded with water, represent stressed, attractive beetle host plants that can be destructively sampled (growers are reluctant to allow sampling of production trees even if heavily infested). Additionally, 25-centimeter-long apple bolts around 3 centimeters in diameter (preferred size for attack by black stem borer), presoaked in ethanol, represent a highly attractive substrate for treatment applications. These are currently deployed in Cornell Campus natural areas previously identified as having large populations of ambrosia beetles. High beetle activity enables rapid elucidation of treatment effects. Anticipation of peak flight times is needed to provide a window of opportunity for successful applications of pesticides (chemical and biological) against ambrosia beetles. Trapping strategies are being tested in and around commercial apple orchards to improve detection of peak activity and to identify pest population centers or “hotspots.” Used in conjunction with orchard surveys, trapping can also pinpoint individual stressed trees that are highly susceptible to attack, enabling growers to spot-treat. High-dose treatments of hotspots or individual trees, as opposed to broadcast treatments at lower doses, can maximize pesticide efficacy while minimizing the amounts of material used. Alcohol-baited traps situated at orchard borders have detected apparent movement of beetles from adjacent wood lots, suggesting potential for elimination of external sources of beetles (for example, via sanitation to remove recently deceased woody materials). Analysis of trap data is resulting in benefits to participating growers. Efforts are also underway to establish a laboratory colony of the granulate ambrosia beetle for investigations of its susceptibility to a broader range of fungal pathogens. This insect is a major pest of woody ornamentals and fruit and nut trees throughout the southeastern U.S. It has recently become a problem also in the northwest and is becoming more prevalent in New York. Objective 2c. Field research by scientists of ARS-Ithaca, New York (in collaboration with researchers of ARS-Hilo, Hawaii and the University of Hawaii, Hilo) aimed at assessing efficacy of spray applications of Beauveria bassiana-based biopesticides against coffee berry borer (CBB) was completed in September 2017. Data collected during four field seasons (2014–2017) in the South Kona and Kau coffee growing districts of Hawaii Island are being analyzed for publication. Specimens of Beauveria collected from more than 8000 adult female CBB found in post-treatment samples were screened as either commercial strain GHA or wild strains, and this information was used to provide accurate estimates of strain GHA efficacy (taking into account the contributions to control by the naturally-occurring strains). Identity of wild strains vs. strain GHA were based on colony morphology in culture and confirmed by genetic testing. Preliminary analysis indicates that activity of the wild strains rarely accounted for greater than 5% mortality in fields treated regularly with strain GHA. However, infection by wild strains could increase to around 20% in these fields under highly favorable conditions (prolonged periods with dense cloud cover, abundant rainfall, high humidity, and moderate temperatures). Under such conditions in untreated fields, even higher rates of infection were observed (up to 42%), but these natural Beauveria outbreaks did not prevent significant crop damage (see accomplishments). Analysis has also revealed that direct infection by spray-applied Beauveria accounted for only 15% mortality of beetles within one week after spray. However, subsequent sporulation of the pathogen on killed beetles resulted in infection of an additional 10% of the CBB (adult female beetles) attacking the crop, even under dry conditions. Each spray application thus produced, on average, 25% mortality of the pest populations sampled within 10 days following a spray. Removal of beetle-infested green berries and raisins (dried berries left in the field after harvest) prior to or very early in the coffee season was found to be a highly effective tactic for beetle control, especially in combination with Beauveria applications. In the absence of constant beetle emergence from the raisin reservoir (where beetle reproduction and development can continue for many months), Beauveria-spray programs resulted in infection of nearly 70% of the beetle population. The combined control tactics limited damage to less than or equal to 10%, even on farms in areas where feral or poorly managed coffee were sources of migrating beetles. Both sanitation (due to labor) and Beauveria spray applications are expensive control options, and if cost limits growers to just one or the other, our findings indicate that sanitation should be recommended. Objective 2d. Studies aimed at genetic and biological characterization of the wild strains of Beauveria bassiana collected from Hawaiian coffee fields has been completed (see accomplishments). Strains representing all major genotypes have been preserved in the ARS Collection of Entomopathogenic Fungal Cultures (ARSEF). Eleven of these strains and several strains from other locations (Puerto Rico and Africa) have been assessed for commercial mass-production potential and bioassayed against CBB to determine their virulence relative to commercial strain GHA. A number of these exhibited virulence and mass-production potential equal to, or greater than, strain GHA. In our field studies, the Hawaiian strains also exhibited a greater capacity for persistence and natural spread in CBB populations than strain GHA, and in view of these numerous positive attributes, we conclude that these fungi are strong candidates for commercial development as biocontrol agents. Additional testing would be required, however, to determine if these fungi possess other key attributes of strain GHA, including activity against a broad range of insect pests and exceptional stability (shelf life) following production and formulation.

1. Biocontrol of fungi against western flower thrips (WFT). Greenhouse growers have indicated that effectiveness of Beauveria- and Metarhizium-based biopesticides is inconsistent under commercial growing conditions. These fungi require high-humidity conditions after application to infect WFT, a possible source of the problem. Scientists of ARS-Ithaca, New York, in collaboration with a visiting scientist from the University of Agriculture, Faisalabad, Pakistan, conducted laboratory tests to determine optimal moisture conditions for fungal efficacy under a range of constant and fluctuating temperatures. We determined that maintaining high humidity for approximately 40 hours following a spray application supported maximum fungal activity. This was observed even when daily cycles included low-temperature periods (simulating nighttime conditions). These results have identified a way for greenhouse pest managers to enhance efficacy and reliability of fungal biocontrol agents.

2. Hidden diversity of insect-pathogenic fungi associated with coffee berry borer (CBB) in Hawaii. CBB is the most important insect pest of coffee in Hawaii. Control is possible but very expensive, and alternative approaches are needed. Researchers from ARS-Ithaca, New York, ARS-Hilo, Hawaii, and the University of Hawaii, Hilo (in collaboration with local coffee farmers) studied the naturally-occurring fungal pathogens of CBB that have been observed to cause over 40% mortality under optimal environmental conditions. DNA sequencing identified nearly all of these fungi as the common insect pathogen Beauveria bassiana but revealed many unique strains or genotypes. These genetic discoveries provided essential information for laboratory studies aimed at assessing potential commercial use of these fungi for CBB control.

3. Natural outbreaks of the insect pathogenic fungus Beauveria bassiana (Bb) do not provide effective control of coffee berry borer (CBB) in Hawaii. Field and laboratory studies have indicated widespread natural occurrence of diverse strains of Bb in Hawaiian coffee. Coffee growers have questioned if recommended control measures can be reduced in fields where these outbreaks occur, saving time and money. Researchers of ARS-Ithaca, New York, ARS-Hilo, Hawaii, and the University of Hawaii, Hilo, studied the outbreaks and found that these wild strains of Bb were unable to hold pest populations below the economic-damage threshold. Our findings have informed growers and extension scientists that natural Bb outbreaks cannot be relied upon to control this difficult pest and that, even when high Bb activity is evident, recommended control measures should not be skipped or delayed.

4. A little-known fungal pathogen of coffee berry borer (CBB) is identified. A fungal pathogen of CBB currently known as Hirsutella eleutheratorum was discovered in Hawaiian coffee fields in 2016. Coffee researchers first observed this fungus infecting CBB in 1993 in Colombia but were unable to grow it in culture. It was nevertheless suggested that the fungus had potential to play an important role in regulation of CBB populations. Researchers from ARS-Ithaca, New York, ARS-Hilo, Hawaii, and the University of Hawaii, Hilo, Hawaii, successfully cultured the Hawaiian strain and conducted a DNA analysis. Results identified it as a member of the citriformis group within the genus Ophiocordyceps but disproved its current classification as the asexual stage of Ophiocordyceps entomorrhiza. In addition to shedding light on the identity of this pathogen, our report describes its exceedingly slow growth in culture and slow spread in CBB populations and informs CBB researchers of its low potential for commercialization as a biocontrol agent.

Review Publications
Castrillo, L., Mayfield, A., Griggs, M., Camp, R., Mudder, B., Taylor, A., Vandenberg, J. 2017. Mortality and reduced brood production in walnut twig beetles, Pityophthorus juglandis (Coleoptera: Curculionidae), following exposure to commercial strains of Beauveria bassiana and Metarhizium brunneum. Biological Control. 114:79-86.
Fisher, J.J., Castrillo, L.A., Donzelli, B., Hajek, A.E. 2017. Starvation and imidacloprid exposure influence immune response by Anoplophora glabripennis (Coleoptera: Cerambycidae) to a fungal pathogen. Journal of Economic Entomology. 110(4):1451-1459.
Wraight, S.P., Galaini-Wraight, S., Howes, R.L., Castrillo, L.A., Carruthers, R.I., Smith, R.H., Matsumoto Brower, T.K., Keith, L.M. 2018. Prevalence of naturally-occurring strains of Beauveria bassiana in populations of coffee berry borer Hypothenemus hampei on Hawaii Island, with observations on coffee plant-H. hampei-B. bassiana interactions. Journal of Invertebrate Pathology.
Morris, E., Stock, S., Castrillo, L.A., Williams, D., Hajek, A. 2018. Characterization of a novel dimorphic Deladenus beddingi n. sp. and its associated woodwasp and fungus. Nematology.
Castrillo, L.A. 2017. The Host Population-Chapter 4. In: Hajek, A.E., Shapiro-Ilan, D.I., editors. Ecology of Invertebrate Diseases. Oxford, UK: John Wiley & Sons Ltd. p. 101-141.