Project : USDA ARS

ARS Home » Southeast Area » Gainesville, Florida » Center for Medical, Agricultural and Veterinary Entomology » Imported Fire Ant and Household Insects Research » Research » Research Project #436291

Research Project: Management of Fire Ants and Other Invasive Ants

Location: Imported Fire Ant and Household Insects Research

2024 Annual Report

Objectives
1. Conduct studies on fire ant biology to develop new and improved surveillance and control strategies as part of an integrated pest management program. 1.1. Develop natural enemies of fire ants as classical biological control agents or biopesticides by characterizing their life cycle, evaluating their effectiveness, determining host specificity, developing methods for production and release, and formulating as biopesticides. 1.2. Determine how irrigation affects fire ant bait efficacy. 1.3. Develop novel biologically-based fire ant control by identifying the behavioral and semiochemical underpinnings of fire ant mating flights and colony establishment. 1.4. Identify key biological processes in fire ant life stages that may be susceptible to disruption. (vacant Molecular Biologist) 1.5. Determine and quantitate fire ant traits that contribute to their ability to survive the harsh conditions associated with accidental transport over long distances and/or establishing and expanding infestations at the invasion front. (vacant Entomologist) 2. Develop new surveillance and control strategies for crazy ants and other invasive pest ants. 2.1. Employ metagenomics techniques and next generation sequencing technologies to discover potential natural enemies of the little fire ant. 2.2. Develop an effective baiting strategy for the control of tawny crazy ants. 2.3. Investigate the homology of pheromone systems that are well understood for S. invicta, but relatively unknown in the little fire ant and the tawny crazy ant. 2.4. Identify key biological processes of little fire ants and/or tawny crazy ants that can be exploited and developed as novel control methods. (vacant Molecular Biologist) 2.5. Determine and quantitate little fire ant, tawny crazy ant, and other ant species traits that contribute to their invasive success, e.g., metabolic rates. (vacant Entomologist)

Approach
1.1 Integration of any new natural agent into a fire ant control program will require the satisfactory completion of studies in host specificity, predicted-efficacy, virulence, mode of action/transmission, formulation/rearing and field release methodologies. 1.2 Water resistant and standard fire ant bait formulations exposed to irrigation will be evaluated for efficacy against fire ant colonies. The effect of bait application methods (piled vs broadcast) on improving bait tolerance to irrigation will also be assessed. 1.3 Behavioral and semiochemical underpinnings of fire ant mating flights and colony establishment will be examined by determining the behavior of alates to pyrazines with olfactometer bioassays and in-flight lek sampling. Male produced tyramides will be further evaluated for physiologicfal functions related to multiple mating and rapid wing loss. 1.4 The approach of this sub-objective will be defined by the scientist filling the vacant Molecular Biologist position. 1.5 The approach of this sub-objective will be defined by the scientist filling the vacant Entomologist position. 2.1 Little fire ants from across the native and introduced ranges will be collected and used as RNA source material to create cDNA expression libraries. Detailed bioinformatics analysis of resulting NGS data will allow us to identify potential microsporidia, fungi, viruses, protists, and non-hymenopteran eukaryotic parasites. Sequence leads will be verified by molecular analysis of little fire ant colonies sampled within and outside the native ranges. 2.2 Consumption and temporal feeding patterns by tawny crazy ants (TCA) on liquid sucrose bait containing a slow-acting toxicant will be compared to bait containing a fast-acting toxicant. Time lapse photography will be used to document temporal feeding patterns over 72 hours. TCA feeding patterns will be used to design liquid bait dispensers such as alginate hydrogel carrier and presented in a compostable dispenser. 2.3 Systematically evaluate exocrine glands in TCA and little fire ant (LFA) workers and queens for phenotypic effects, e.g. attraction, repellency, alarm, and recruitment using behavioral bioassays. Attraction will be investigated first, using a Y-tube olfactometer bioassay to guide the isolation of active compounds. Attractants can enhance baits and improve monitoring systems. 2.4 The approach of this sub-objective will be defined by the scientist filling the vacant Molecular Biologist position. 2.5 The approach of this sub-objective will be defined by the scientist filling the vacant Entomologist position.

Progress Report
This is the final progress report for project 6036-32000-051-000D which terminates on 09/02/2024. (Objective 1) The development and commercialization of a red, black, and hybrid imported fire ant detection device was completed and has been available to APHIS, stakeholders, and the public from Agdia, Inc. as InvictDetect™ Plus. The device, a laminar flow immunoassay (analogous to a pregnancy test) provides a much-needed tool for regulatory agencies in the USA and other countries to enforce quarantine protocols to limit the spread of these invasive ants. This progress significantly improves surveillance of these invasive fire ant species through a single, rapid, on-site test. (Objective 1.1) A fire ant specific virus was released and established in California and Florida for the biological control of red imported fire ants (RIFA) to improve fire ant IPM. Field inoculations of fire ant nests with the virus Invictavirus solenopsae (SINV-3) in Florida caused reductions in the size and number of fire ant nests, as well as persistence and spread of the virus to adjacent uninoculated colonies, which extended the control. SINV-3 provides an additional natural control agent against fire ants with an advantage over traditional insecticides of no known detrimental ecological impacts, host specificity, and sustainability. In addition to Florida, augmentative releases of the virus were successful in California. ARS researchers at Gainesville, Florida completed research to determine the mode of action of the virus, Invictavirus solenopsae, in RIFA. While Invictavirus solenopsae has been shown to cause significant mortality among all stages of its RIFA host, the mechanism by which it does so has remained unresolved. Studies showed that virus infection of the ant causes damage to the midgut cell lining of worker ants and alters their feeding and foraging behavior. The worker ants stop acquiring food for the colony causing starvation, which leads to mass mortality and queen wasting. Fire ants were collected from 360 nests along a transect from south Florida to north Georgia. and from 66 sites in the panhandle of north Florida (collaborating with Florida A&M University). DNA extractions from these samples were used to survey for the microsporidian fire ant pathogen Kneallhazia solenopsae. Future sequencing data will deepen our understanding of the genetic diversity of K. solenopsae in the USA. (Objective 1.2) Fire ants can be efficiently controlled by fire ant baits which typically contain corn grit, an ingredient that allows the baits to be easily applied and foraged by ants. Traditionally it is thought that corn grit exposed to rain or irrigation absorbs moisture and compromises bait efficacy because ants do not feed on wet bait. Water-resistant fire ant baits have been developed that either replace or modify the corn grit. Research conducted with collaborators from APHIS, and the Coachella Valley Mosquito and Vector Control District in California determined that both standard commercial fire ant bait and water-resistant baits that have been soaked in water in the laboratory, as well as exposed to sprinkler irrigation in landscapes, can still effectively control fire ants. Because water-resistant fire ant baits are not readily available in the U.S., knowledge that standard fire ant baits can withstand sprinkler irrigation should allow land managers more flexibility in scheduling bait treatments. Nursery stock with root balls wrapped in burlap that are shipped outside the Federal Imported Fire Ant (IFA) Quarantine must be free of fire ants. Water-resistant spatter and granular ant baits applied to irrigated, balled and burlappd plants reduced fire ant populations, but inconsistently eliminated fire ant colonies due to intermittent fire ant foraging relative to bait palatability and cool weather. These bait treatments did not meet IFA quarantine standards. An effective alternative of spraying non-repellent, contact insecticides to the exterior of infested root balls demonstrated a consistent elimination of fire ant colonies. In addition, this treatment also prevented root balls from being infested by migrating fire ant colonies for 4–6 months. It is hypothesized that the nonrepellency facilitated greater ant contact with the insecticide, resulting in better control than traditional treatments. This discovery has led to further cooperative research with the nursery industry through collaborations with Tennessee State University and APHIS. (Objective 1.3) Initially, fire ant mating flights were used to determine how newly-mated queens quickly overcame the dealation inhibitory effects imposed on them by queen pheromones. This research quickly changed when it was published that male fire ants produce tyramides. Over the next several years it was determined that males transfer the tyramides to female sexuals during mating. The female sexuals release a specific enzyme during the mating process that hydrolyzes tyramides to the biogenic amine, tyramine. Metabolomic studies determined that newly-mated fire ant queens have highly elevated amounts of tyramine in their gasters, head and thorax. Tyramine injected into mature female sexuals 1) precociously lost their wings (dealate); 2) started reproductive development; and 3) produced queen pheromone (a worker attractant). This sequence of events was published in Nature Communications. The use of these findings for fire ant control consisted of feeding tyramine to colonies that had female sexuals, such that they would dealate and compete with the real queen, resulting in decreased egg production. Notably, the treatment with tyramine resulted in significant worker mortality with lab colonies. A patent application was submitted, and we engaged a commercialization partner with whom we had Phase 1 and 2 SBIR grants and associated CRADAs. Field studies clearly showed that field (wild) fire ants were repelled by bait formulations with tyramine. An NSF SBIR phase 1 grant allowed us to investigate methods to overcome the negative taste of tyramine using taste masking methods already developed to mask the bad taste of medications associated with human pharmaceuticals. This research resulted in a formulation that is readily consumed by wild fire ants. Field evaluations are being developed now. (Objective 2.) Collaborating with researchers from Ecuador, the invasive, tropical fire ant, Solenopsis geminata, were collected from >400 nests in the Galapagos Islands and continental Ecuador. These ant samples will be used to track routes of introduction of this invasive ant into the Galapagos Islands using both genetic and chemical markers; plus they will be screened for microorganisms with biocontrol potential. (Objective 2.1) Employing metagenomics and next generation sequencing technologies has led to the discovery of the first viruses from the invasive little fire ant, Wasmannia auropunctata. This ant is one of the most destructive invasive ants worldwide. Seven viruses were discovered in the little fire ant from its native range, which offer potential classical biological control agents against this pest ant in the USA and other regions globally. (Objective 2.2) Efforts to control overwhelming numbers of tawny crazy ants (TCA) in buildings and landscapes often rely on broadcast applications of residual, contact insecticides and granular ant baits. However, suppression often is temporary and inadequate. Our research has shown that a fast-acting liquid bait containing dinotefuran results in significant and visually perceptible reductions in TCA, but its effects were localized, and areas became reinfested within 2-4 weeks after baiting stopped. To extend bait accessibility, a laboratory study indicated that TCA readily fed on alginate hydrogel carrier containing liquid ant bait that was dispensed in a compostable station. Field deployment of these stations revealed hydrogel desiccation and limited ant presence at the stations; thus more research is needed. To improve baiting efficacy, comparisons of fast and slow-acting bait toxicants (dinotefuran and disodium octaborate tetrahydrate, respectively) in sucrose solution demonstrated significant reductions in TCA nesting and foraging activity with each bait type. However, once baits were not replenished, TCA populations began to return. Strategic bait placements where slow-acting bait was placed along the field plot perimeters to allow bait distribution beyond the plots, while the fast-acting bait was centered within the plots to quickly suppress TCA. This baiting strategy resulted in significantly less TCA nesting and foraging within the plots. (Objective 2.3) The response of little fire ant (LFA) workers to trail pheromone chemicals deposited on various substrates was investigated. While the pheromones responsible for trail formation were not identified, the groundwork of developing a suitable bioassay was completed and published. The LFA alarm pheromone was previously identified as a pyrazine derivative with multiple substituents on the pyrazine ring. This is like the tri-substituted pyrazine isolated and identified for the red imported fire ant (RIFA). In the case of RIFA, and likely to be similar for LFA, the alarm pheromone acts to both increase the worker ant activity and attract the worker ants to the source of the pyrazine. Therefore, applications can be an efficient attractant for traps or baits. In addition, it was determined that extracts of LFA worker abdomens elicited rapid movement in previously quiescent workers in a fully functional LFA colony. The identification of the responsible semiochemicals will be a future target.

Accomplishments
1. Mode of action of a fire ant killing virus. Invictavirus solenopsae (previously called Solenopsis invicta virus 3) is a virus specific to fire ants that is an effective natural control agent for fire ants in the United States. Numerous stakeholders (APHIS, pest controllers, public) are interested in this research because the virus provides a specific, non-toxic, sustainable method of controlling invasive fire ants. However, the mechanism of action of the virus is not completely understood. Scientists in Gainesville, Florida determined that Invictavirus solenopsae infection can be transmitted from immature stages to adults, possibly through the adult care of the immature ants. Furthermore, the adult ants stop acquiring food for the colony, causing starvation, which leads to mass mortality of the immature ants and queen wasting. These results advance the understanding and development of this virus as a biological control agent for fire ants.

2. New biodegradable, species-specific peptides inactivate fire ant colonies. It has been over 25 years since the last fire ant bait control product was commercialized. Neuropeptides such as Pheromone Biosynthesis Activating Neuropeptides (PBAN) activate receptors to initiate essential biochemical reactions in insects. ARS scientists in Gainesville, Florida and Corvalis, Oregon used their patented Receptor-interference technology to specifically isolate peptides that strongly bind to and inactivate fire ant PBAN receptors. Selected peptides fed to fire ant colonies resulted in high worker mortality and death or sterilization of the queen. This proof of concept of the Receptor–interference technology may lead to the development of a new fire ant bait that is biodegradable and specifically targets fire ants.

3. An inexpensive fire ant bait formulation. Commercially available fire ant baits are typically sold to households and other high-value markets. However, these products are too expensive for farmers and ranchers who need fire ant control over large acreages. An ARS scientist in Gainesville, Florida and a CRADA partner discovered inexpensive fire ant toxicants. They determined that encapsulation of the toxicants allowed passage into the fire ant gut resulting in high worker ant mortality and queen death. This novel bait formulation is expected to be less expensive than currently available fire ant baits and may provide an economical fire ant control option accessible to farmers and ranchers.

4. Tawny crazy ant control with baits. The tawny crazy ant (TCA) is an invasive ant from South America that is established in the Gulf Coast states from Florida to Texas, plus Georgia. It develops overwhelming populations that can inundate urban, rural, and natural landscapes. Attempts to control TCA populations often rely on excessive applications of residual, contact insecticides over entire landscapes, which is often temporary and inadequate. Researchers from Gainesville, Florida have demonstrated that a fast-acting liquid bait containing dinotefuran resulted in significant and visually perceptible reductions in TCA, but its effects were localized, and areas became reinfested within 2-4 weeks after baiting was discontinued. Comparison of TCA foraging intensity in the field on the fast-acting bait and a slow-acting, boric acid based, liquid bait indicated sustained and significantly more consumption of the slow-acting bait. In addition, significant reductions in tawny crazy ant nesting and foraging activity occurred with both types of baits. A baiting strategy utilizing a combination of both fast and slow-acting liquid baits has resulted in sustained and more expansive control, which can curtail excessive insecticide applications often used against this invasive ant.

U.S. DEPARTMENT OF AGRICULTURE

Imported Fire Ant and Household Insects Research: Gainesville, FL