Research Project: Exotic Whitefly and Subtropical Invasive Pests of Vegetables and Ornamental Plants

Location: Subtropical Insects and Horticulture Research

2023 Annual Report

Objectives
Objective 1: Investigate the interactions of subtropical invasive pests of vegetables and ornamental crops (e.g., tomato, pepper, floriculture) with their natural enemies and use this information to develop biocontrol-based banker plant systems for environmentally sound pest management. [NP304, C3, PS3A, 3B and 3C] Sub-objective 1a: Expand the utility of the papaya banker plant system by incorporating predators and/or entomopathogenic fungi simultaneously or in combination with the parasitoid. Sub-objective 1b: Build a better banker plant through biotechnology by genetically engineering papaya to contain attributes that make it more suitable for banker plant use: i.e. nonflowering, papaya ringspot virus and powdery mildew resistant. Sub-objective 1c: Develop a mealybug banker plant system using ornamental muhly grass (Muhlenbergia capillaris), the native mealybug (Stemmatomerinx acircula), and predatory beetles (Diomus austrinus, Cryptolaemus montrouzieri) for control of pest mealybugs. Objective 2: Investigate structural, physiological, molecular, and chemical aspects of whiteflies and identify inhibitor strategies/molecules such as but not limited to feeding disruptors and peptide inhibitors of disease transmission that can be used in the development of novel interdiction strategies envisioned to work either through production of transgenic plants or application of chemical treatments that block feeding/disease transmission. [NP304, C3, PS3A, 3B and 3C] Sub-objective 2a: Identify interdiction molecules that can be expressed in transgenic or Plant-Host Activated-Cell Transplantation (PHACT) adapted plants for controlling hemipteran insects and their transmitted diseases. Sub-objective 2b: Development of transgenic vegetables and ornamentals with increased resistance to hemipteran pest insects and/or their transmitted diseases Sub-objective 2c: Plant-Host Activated-Cell Transplantation (PHACT) as a strategy to induce plant resistance to hemipteran insects and their transmitted diseases.

Approach
This project focuses on two control strategies for whitefly and subtropical invasive. The first approach will be the development of banker plant systems for whitefly and mealybug management that will be used to establish, augment and increase the numbers of natural enemies in vegetable and ornamental production. We will expand the utility of the papaya banker plant system to include general predators and/or entomopathogenic fungi to enhance the pest control capability of the system by increasing the number and diversity of pests controlled. We will construct a nonflowering papaya banker plant through biotechnology that is both Papaya ringspot virus and powdery mildew resistant to increase the utility of the banker plant system. Mealybug banker plant systems will be developed to provide viable biologically control alternatives for environmentally sound pest management for nursery, greenhouse ornamental and vegetable growers. The second approach focuses on the development of a biological interdiction strategy through identification of entomotoxic biomolecules (primarily peptides) that target the pest insect or the transmitted pathogen and delivering these molecules either through transgenic expression or through a novel Plant-Host Activated-Cell Transplantation (PHACT) organoid-biofactory strategy. The outcomes of this research range from development of improved and sustainable IPM strategies employing banker plant systems to identification of new biologically-based control strategies incorporating either resistant transgenic plants or deployment of an engineered plant organoid biofactory system for hemipteran pest insect control.

Progress Report
Two greenhouse trials arranged in a randomized complete block design (RCBD) with four treatments and 4 replications were conducted, by ARS scientists in Fort Pierce, Florida, to determine the effectiveness of pallidus beetle to control Bemisia tabaci MEAM1 populations applied either directly to poinsettia or by a papaya banker plant system compared to the insecticide grower standard at the high (first greenhouse trial) and low rate (second greenhouse trial) and the untreated control. Preventative fungicide for powdery mildew was applied to the banker plants prior to infestation with papaya whitefly and the predatory beetle. No powdery mildew was detected on the banker plants in either trial. Adult pest whitefly populations in the untreated control in both trials reached 525 per leaf and resulted in plant death. Insecticide control was excellent for the duration (13 weeks) of trial one at the high rate and control broke starting in week 7 of the second trial at the low insecticide rate. Both beetle treatments provided significantly better control of all stages of the pest whitefly compared to the untreated control and were not significantly different from the insecticide treatment for most weekly evaluations. Beetles from the papaya banker plant dispersed readily to the cash crop and provided better control of Bemisia overtime compared to direct release. Experiments to test the efficacy of fungicides sprayed on papaya banker plant systems (papaya whitefly, predatory beetle and/or parasitoid) with powdery mildew to control the disease and not harm the components of the system were not conducted. Under the contingencies of 1b, we projected to apply curative applications of fungicides against powdery mildew on banker plants in the presence of whiteflies and beetles on papaya banker plants but did not get powdery mildew infection on any of the experimental banker plants at the level we can conduct this experiment. We will collaborate with plant pathologists to in-vitro culture powdery mildew in the laboratory so banker plants can be uniformly infected at various levels (low, medium, high) for experiments in FY2024. Finished laboratory trials determining the temperature-dependent development of Stemmatomerinx acircula at a range of 10-35°C. Host specificity tests of S. acircula on economically important crops were also finished and these studies will be combined into a single paper. Data shows that S. acircula is highly specific to just a few closely related Muhlenbergia species and can hatch at temperatures up to 35°C, but first instar do not survive long. These studies suggest a S. acircula banker plant system may be viable in many greenhouse cropping systems in the Southeastern US. Manuscript in preparation. Finished conducting greenhouse studies using a S. acircula muhly grass banker plant system with the predatory beetle Diomus austrinus for managing pest populations of the highly invasive Madeira mealybug (Phenacoccus madeirensis) on Coleus as the ornamental cash crop. While control is eventually achieved over many weeks, it was determined that this release strategy was not a viable economic management option for Madeira mealybug. Crop damage thresholds reached unacceptable levels before beetles could reduce mealybug populations. Diomus appears to prefer mealybug eggs so need something that would attack later stages. Alternative options are being considered, such as adding mealybug parasitoids in conjunction with the predatory beetles. Purchased Cryptolaemus montrouzieri (mealybug destroyer) from different biological control companies (BioB, IPM Labs, RINCON bitova) with multiple releases from each source as shipments arrived overtime. Each Cryptolaemus source company had a greenhouse and were evaluated on open populations of Citrus mealybug infesting 3 ornamental host plants (Croton, poinsettia, celosia). None of the shipments from the various biological control companies were able to establish on any of the three ornamental host plants screened. A single cage out of 3 had reproduction of the mealybug destroyer, but not enough to establish a population despite multiple releases. Regardless of the source of Cryptolaemus, all shipments had problems with quality control, predator establishment, dispersal to the pest plus these predators are too big and potential clientele (nursery and greenhouse growers) do not like them. Preliminary work indicated that Agrobacterium tumefaciens produced galls on papaya induced plant host defenses that had a strong negative impact on whitefly fecundity. As a result, a screen of Agrobacterium genotypes was performed to evaluate the ability to form galls of desirable type that could be used in a whitefly biocontrol strategy. Three strains were identified for further evaluation and whitefly experiments were initiated to measure the effect of galls produced by these Agrobacterium strains on whitefly fecundity. Combinations of plants and agrobacterium strain interactions were evaluated to identify combinations that would support the use of agrobacterium induced galls as harvestable Biofactories. Two plant types, Ficus and Euonymus were identified as being able to produce numerous large galls with specific Agrobacterium strains and these were selected for further evaluation to produce desirable biomolecules that could be used in pest insect and pathogen control in horticultural crops. Finished conducting laboratory trials comparing the suitability of a diet of decapsulated cysts of the brine shrimp Artemia franciscana to larvae of Echinothrips americanus for development and reproduction of Franklinothrips vespiformis. The effects of alternate diet on predation of E. americanus larvae was also examined. Data collection is complete, and analysis is underway. Preliminary data analyses suggest that egg to adult development is similar between diets, but adult longevity is significantly longer in individuals fed on Artemia cysts (45 vs 38 days). The presence of Artemia cysts also significantly reduced predation of E. americanus larvae (12 vs 7 larvae/day). Manuscript preparation is ongoing. Finished laboratory trials to determine the functional response of the pantropical predatory thrips (Franklinothrips vespiformis) on whitefly (Bemisia tabaci) to assess its impact as a predator of whiteflies. Our results showed that F. vespiformis was a very effective predator, with adults consuming up to 110 eggs and 30 nymphs per day. Data analysis is complete, and the manuscript is ready to be sent to reviewers. Future work will assess various release strategies for F. vespiformis for B. tabaci management and compare the use of F. vespiformis against chemical control and other commonly used biological control agents. Extensive Thrips parvispinus surveys of box cutter stores were conducted in Palm Beach County and were detected in 13 of the 14 stores surveyed, primarily on gardenia, Dipladenia and pepper transplants. Surveys were extended to north Florida counties (9) including several that border Georgia and the pest was found in all stores surveyed bringing the positive county tally in Florida to 26 counties. This pest was detected in vegetable fields for the first time in the United States which resulted in a $1.3 million dollar crop loss for that vegetable grower. Samples received from three new state records were submitted and confirmed by The Animal and Plant Health Inspection Service (APHIS) for Georgia, South and North Carolina. Suspect samples from Ohio are being processed and submitted to APHIS for confirmation. Developed a website and field guide containing information about early diagnostics, field scouting, biology, and management options for T. parvispinus. Conducted genetic analysis of its global populations and revealed 109 sequences of its COI gene submitted to GenBank divided into 13 haplotypes. One of these 13 haplotypes is shared across continents and has invaded Florida. Distributed field guides among stakeholders through the University of Florida extension offices. Conducted hands-on diagnostic training workshops, webinars and presented multiple presentations (Jenson Beach Garden Show, Working Groups, Horticulture Research Institute webinar, IFAS webinars, Citrus Show 2023) and wrote outreach articles for GrowerTalks. Manuscript in preparation. In support of the USDA/ARS Innovation Fund Award (Round 14): the unanswered question is whether the potassium in 10% KOH or its pH turns hemolymph green. Tested alternative solutions with different pHs in the laboratory and discovered that pH, not potassium, was turning hemolymph green, and 5% of Pure Lye solution (sodium hydroxide, NaOH) with a pH of 14 turns Nipaecoccus viridis hemolymph green in less than 1 minute. To our knowledge, the hemolymph of three more species (Amonostherium hymenocleae, Amonostherium lichtensioides, and Paracoccus hakeae) from other genera of mealybugs can also turn green in 10% KOH. Managed to get their samples from Arizona, California, and Hawaii. Tested with our alternative solution (5% Lyle) as positive controls. Paracoccus marginatus previously known to not turn green was used as a negative control in our tests. Confirmed that 5% NaOH with pH 14 also turns other species green. Manuscript submitted. Analyzed Wolbachia surface protein (wsp) gene sequences along with all possible available datasets of wsp from Bemisia cryptic species to study the evolutionary history of this gene and Wolbachia. Identified 30 genotypes, 22 of which belonged to the genetic identity wBt1 that was predominant and occurred in members of B. tabaci complex found in Asia, Africa, Australia, and Europe. Found evidence of identical wsp genotypes between indigenous B. tabaci and members of the complex that have recently invaded China. Studies are exploring the potential routes of horizontal transmission of Wolbachia globally in the Bemisia species complex. Manuscript in preparation.

Accomplishments
1. Distribution of Scirtothrips dorsalis cryptic species complex in the United States and reproductive host assessment of its dominant member. Chilli thrips, Scirtothrips dorsalis Hood, is a cryptic species complex (group of morphologically indistinguishable species) of at least nine distinct species, two (South Asia 1 and East Asia 1) of which have been confirmed damaging >50 plant taxa in the United States. To develop a knowledge-based management program for this pest, a nationwide survey for chilli thrips was conducted by ARS scientists in Fort Pierce, Florida. Of the 101 thrips populations received across the United States between 2015 and 2021, 71 populations were confirmed as S. dorsalis, with ~25% further identified as East Asia 1 from temperate Japan, compared with ~41% identified as South Asia 1 (India), suggesting the latter species is more prevalent in the US. East Asia 1 was found almost exclusively in samples collected from Hydrangea sp. (95%) in Massachusetts, New York, and Pennsylvania, indicating geographic range expansion in the Northeast and successful overwintering in areas with hard freezes contradicting previous CAPS surveys which forecast this species would not survive in northern regions. While assessing the host range of South Asia 1 (dominant species), 62 plant taxa were evaluated under greenhouse conditions. Among these, 40 feeding and reproductive hosts were confirmed in Florida with 18 new reproductive and 3 feeding hosts not previously reported in the literature. This study highlights why correctly identifying and tracing an invasive pest’s origin is important to develop a robust management practice and delimit its geographical distribution. Specifically in the case of cryptic species, misidentification can lead to the misapplication of management practices, resulting in wasted money, resources, and time.

2. Biology, distribution, field guide, host plants, and current management tools of lebbeck mealybug. Lebbeck or hibiscus mealybug (LM), Nipaecoccus viridis, is an invasive, highly polyphagous agricultural pest that attacks food, forage, ornamental, and fiber crops, especially citrus. In citrus-growing areas of Jordan, where LM had been a pest before the introduction of biological controls, it caused such extensive damage that groves were burned to eradicate it. Though global invasion of LM started over a hundred years ago, and LM has been reported from 67 countries from 5 continents (Africa, Australia, Asia, Europe, and North America), Oceania, and the Caribbeans feeding on 227 host plant species from 148 genera of 53 host plant families, its diagnostics and management tools were still not available. Our studies provide its biology, distribution, field guide, host plants, and current management options. A field guide presented in this study will help detect the pest early and prevent future introductions to new locations and plant hosts. ARS scientists in Fort Pierce, Florida, organized the mealybugs symposium with colleagues at the University of Florida in the Southeastern Branch Meeting of the Entomological Society of America held in Little Rock, Arkansas, and presented this field guide. In addition, we collaborated with colleagues at the University of Florida, organized a mealybugs workshop for nursery stakeholders, and demonstrated the field guide during the workshop in front of over 45 participants. We anticipate that the information will help slow its spread in the U.S.

3. Documenting predatory ladybird beetles’ diversity and their prey whitefly species in Hawaii. Ladybird beetles (LB) are an important group of insect predators that prey upon many pests, including aphids, scale insects, mealybugs, and whiteflies, in crop fields, orchards, gardens, urban landscapes, and greenhouses. Several species of LB have been purposefully and inadvertently introduced into the Hawaiian Archipelago for over 100 years. Some have a tremendous potential to be used in the biological control of invasive pests, including whiteflies. However, studying their predatory potential would not be possible without documenting the species of LB and their prey. Here, ARS scientists in Fort Pierce, Florida, updated the list of LB in Hawaii with two new state records, 17 new island records associated with 13 species, and various prey records, including five whitefly species. The update includes new geographic distribution records from the eight main and Northwestern Islands. The new records demonstrate the value of ongoing sampling and regular examination of undetermined collections material. These findings will assist stakeholders who work with insect management in Hawaii, including pest management practitioners, agriculturalists, conservationists, and biodiversity specialists.

4. Parasitoid vector bacteria among whiteflies: unintended aspect of whitefly biological control. Parasitic wasps play a vital role in the sustainable control of whitefly worldwide. Encarsia formosa is one of the most commonly used parasitic wasps in whitefly biological control. Whitefly harbors one of the most diverse bacterial endosymbiont communities, including Arsenophonus, Cardinium, Hamiltonella, Rickettsia, and Wolbachia. Among them, Rickettsia can modify host biology and manipulate the host’s reproduction to enhance its spread in whitefly. The ecological interactions between parasitic wasps and whiteflies have long been proposed to mediate the horizontal transfer of bacterial endosymbionts within the whiteflies but have not been demonstrated in E. formosa and whitefly. Our study reveals that E. formosa can pick Rickettsia from infected whitefly during parasitism, retain it in its body for at least 48 hours, and deliver it to other uninfected whiteflies. As a result of this transinfection, Rickettsia could modify the host biology and enhance its pestiferous nature. These results question the use of E. formosa in the commercial biological control of whitefly and point out an unintended impact of parasitic wasps in whitefly biological control.

Review Publications
Olabiyi, D., Middleton, E., Ahmed, M.Z., Osborne, L., Mckenzie, C.L., Diepenbrock, L.M. 2023. Hibiscus mealybug (hemiptera: pseudococcidae)- biology, field guide, North American host plants, and current management prospects. Journal of Integrated Pest Management. 14 (1). https://doi.org/10.1093/jipm/pmac029.
Liu, Y., He, Z., Wen, Q., Peng, J., Zhou, Y., Mandour, N., Mckenzie, C.L., Ahmed, M.Z., Bao-Li, Q. 2023. Parasitoid-mediated horizontal transmission of Rickettsia between whiteflies. Frontiers in Cellular and Infection Microbiology. https://doi.org/10.3389/fcimb.2022.1077494.
Díaz-Sánchez, J.L., Serna, F., Ahmed, M.Z. 2022. New records of whiteflies (Hemiptera, Aleyrodidae) and their host plants from Colombia. Biodiversity Data Journal. 18(4):889-896. https://doi.org/10.15560/18.4.889.
Hesler, L.S., Perreira, W.D., Matsunaga, J.N., Yee, D.A., Ahmed, M.Z., Beckendorf, E.A. 2023. New state, island and prey records from Hawai'i, U.S.A., and a new country record from Laos for lady beetles (Coleoptera: Coccinellidae). The Pan-Pacific Entomologist. 99(2):128–141. https://doi.org/10.3956/2022-99.2.128.
Deeter, L.A., Ahmed, M.Z. 2023. The records of Nipaecoccus viridis (Newstead) (Hemiptera: Pseudococcidae) deposited in the Florida State Collection of Arthropods. Insecta Mundi. 0995: 1–8.
Kumar, V., Xiao, Y., Borden, M.A., Ahmed, M.Z., Mckenzie, C.L., Osborne, L. 2023. Distribution of Scirtothrips dorsalis cryptic species complex in the United States and true host assessment of its dominant member. Journal of Economic Entomology. 2023. https://doi.org/10.1093/jee/toad138.
Ahmed, M.Z., Dorado, C., Ellenrieder, N.V., Quinn, N., Roda, A., Schoeller, E.N., Mckenzie, C.L., Osborne, L.S., Diepenbrock, L.M. 2023. Development of a species-level field diagnostic kit for Nipaecoccus viridis (Newstead) (Hemiptera: Pseudococcidae), an invasive and regulatory pest in the United States. Journal of Applied Entomology. 1-9. https://doi.org/10.1111/jen.13177.