Faith M. Oi

RNA interference in the termite Reticulitermes flavipes through ingestion of double-stranded RNA

RNA interference (RNAi) represents a breakthrough technology for conducting functional genomics research in non-model organisms and for the highly targeted control of insect pests. This study investigated RNAi via voluntary feeding in the economically important pest termite, Reticulitermes flavipes. We used a high-dose double-stranded (ds) RNA feeding approach to silence two termite genes: one encoding an endogenous digestive cellulase enzyme and the other a caste-regulatory hexamerin storage protein. Contrary to results from previous low-dose studies that examined injection-based RNAi, high-dose silencing of either gene through dsRNA feeding led to significantly reduced group fitness and mortality. Hexamerin silencing in combination with ectopic juvenile hormone treatments additionally led to lethal molting impacts and increased differentiation of presoldier caste phenotypes (a phenotype that is not capable of feeding). These results provide the first examples of insecticidal effects from dsRNA feeding in a termite. Additionally, these results validate a high-throughput bioassay approach for use in (i) termite functional genomics research, and (ii) characterizing target sites of conventional and novel RNAi-based termiticides.

Juvenile hormone and colony conditions differentially influence cytochrome P450 gene expression in the termite Reticulitermes flavipes.

In lower termites, the worker caste is a totipotent immature stage that is capable of differentiating into other adult caste phenotypes. We investigated the diversity of family 4 cytochrome P450 (CYP4) genes in Reticulitermes flavipes workers, with the specific goal of identifying P450s potentially involved in regulating caste differentiation. Seven novel CYP4 genes were identified. Quantitative real‐time PCR revealed the tissue distribution of expression for the seven CYP4s, as well as temporal expression changes in workers in association with a release from colony influences and during juvenile hormone (JH)‐induced soldier caste differentiation. Several fat‐body‐related CYP4 genes were differentially expressed after JH treatment. Still other genes changed expression in association with removal from colony influences, suggesting that primer pheromones and/or other colony influences impact their expression. These findings add to a growing database of candidate termite caste‐regulatory genes, and provide explicit evidence that colony factors influence termite gene expression.

Speed of Efficacy and Delayed Toxicity Characteristics of Fast-Acting Fire Ant (Hymenoptera: Formicidae) Baits

Efficacy and speed of action of fire ant (Hymenoptera: Formicidae) baits that claim fast control of colonies were compared with a standard bait. More than 85% of red imported fire ant, Solenopsis invicta Buren, laboratory colonies provided bait containing the active ingredient indoxacarb died within 3 d, and all colonies were dead in 6 d. Standard bait containing hydramethylnon resulted in death of 60% of the colonies in 9 d. Bait containing spinosad did not cause colony death. Under field conditions, one-half of the areas treated with the indoxacarb bait did not have any active fire ant nests within 3 d, whereas 11 d was needed to reach the same level of control with the hydramethylnon bait. Spinosad had a maximum of 17% of the treated areas without nests after 3 d. The delay in death of S. invicta adults treated in the laboratory with the indoxacarb and spinosad baits was shorter than the standard hydramethylnon bait, which had mortality similar to the traditional delayed toxicity criterion of < 15% mortality after 24 h and > 89% mortality over the test period. Indoxacarb caused mortality of 57% at 24 h and 100% at 48 h; however, visual symptoms of toxicity were not readily observed for at least 8 h before the abrupt increase in death. Spinosad caused 96% mortality by 24 h, and initial mortality became apparent at 4 h. Time required for death of 15% of a treated population (LT15) of spinosad, indoxacarb, and hydramethylnon was 3, 9, and 16 h, respectively. Delayed toxicity characteristics of the fast-acting indoxacarb bait may be useful for the development of other fast-acting ant baits.

Detection of Permethrin Resistance and Fipronil Tolerance in Rhipicephalus sanguineus (Acari: Ixodidae) in the United States

ABSTRACT Permethrin is a commonly used acaricide for tick control on domestic animals and in residential environments, while fipronil use is restricted to on-animal treatment. Following widespread reports of permethrin and fipronil application failures to control indoor infestations of Rhipicephalus sanguineus (Latreille), the brown dog tick, 31 tick populations were obtained from Florida and Texas for acaricide resistance screening. These field-collected ticks from kennels and residential facilities were challenged with technical grade permethrin and fipronil to create dose response curves that were compared with an acaricide-susceptible strain. Permethrin resistance was successfully screened in nine populations, all of which were resistant or highly resistant. Tick susceptibility to fipronil was conducted on four populations, which were found to be tolerant, with resistance ratios below 10. This is the first documentation of R. sanguineus permethrin resistance and fipronil tolerance in the United States. Potential causes of resistance development and recommendations on future brown dog tick control management plans are discussed.

Establishing the discriminating concentration for permethrin and fipronil resistance in Rhipicephalus sanguineus (Latreille) (Acari: Ixodidae), the brown dog tick

BACKGROUND Rhipicephalus sanguineus (Latreille), the brown dog tick, is a veterinary canine and urban pest. These ticks have been found to develop resistance and tolerance to the two commonly used acaricides permethrin and fipronil respectively. We have developed a discriminating concentration that can be used for rapid detection of permethrin and fipronil resistance in brown dog tick populations. The availability of a discriminating concentration for the brown dog tick provides an inexpensive and rapid resistance diagnostic technique that can be used to guide tick management plans for companion animals and aid in the selection of environmental treatment options. RESULTS Establishing the discriminating concentration for permethrin and fipronil in brown dog ticks enables a resistant:susceptible screening. For permethrin the discriminating concentration was set at 0.19%, and for fipronil at 0.15%. Three additional diagnostic concentrations were chosen to evaluate resistance levels when larval tick numbers were available for screening. CONCLUSION Future tick submissions from residences and kennel facilities can be subjected to a single chemical concentration to diagnose resistance, which minimizes time, costs and tick rearing requirements and guides effective control plans. With the standardized use of larval ticks, a client-submission quantity of ideally five engorged females would provide sufficient larval numbers to utilize this technique.

Discriminating concentration establishment for permethrin and fipronil resistance in Rhipicephalus sanguineus (Latreille) (Acari: Ixodidae), the brown dog tick

BACKGROUND Rhipicephalus sanguineus (Latreille), the brown dog tick, is a veterinary canine and urban pest. These ticks have been found to develop resistance and tolerance to the two commonly used acaricides permethrin and fipronil respectively. We have developed a discriminating concentration that can be used for rapid detection of permethrin and fipronil resistance in brown dog tick populations. The availability of a discriminating concentration for the brown dog tick provides an inexpensive and rapid resistance diagnostic technique that can be used to guide tick management plans for companion animals and aid in the selection of environmental treatment options. RESULTS Establishing the discriminating concentration for permethrin and fipronil in brown dog ticks enables a resistant:susceptible screening. For permethrin the discriminating concentration was set at 0.19%, and for fipronil at 0.15%. Three additional diagnostic concentrations were chosen to evaluate resistance levels when larval tick numbers were available for screening. CONCLUSION Future tick submissions from residences and kennel facilities can be subjected to a single chemical concentration to diagnose resistance, which minimizes time, costs and tick rearing requirements and guides effective control plans. With the standardized use of larval ticks, a client-submission quantity of ideally five engorged females would provide sufficient larval numbers to utilize this technique.

Insecticides for Suppression of Nylanderia fulva

Nylanderia fulva (Mayr) is an invasive ant that is a serious pest in the southern United States. Pest control operators and homeowners are challenged to manage pest populations below acceptable thresholds. Contact and bait insecticides are key components of an Integrated Pest Management (IPM) strategy, however, little is known about their efficacy. In repellency and efficacy bioassays, N. fulva were not completely repelled by any insecticide tested, although fewer ants crossed a surface treated with Temprid®. Few insecticides provided rapid control. Termidor® and Temprid® were the best performing with mean mortality of 100% in 13.4 and 19.0 days, respectively. In no-choice bait acceptance studies, it was shown that N. fulva generally had greater acceptance of carbohydrate-based ant baits (Advion®, InTiceTM (gel), and InTiceTM (granular)). However, mortality was low for the InTiceTM baits in a 7-day bioassay. Maxforce® Ant Killer Bait Gel and Advance® 375A in the spring and Maxforce® Complete in the summer and fall required the fewest days to reach 100% mortality. Bait active ingredients that resulted in the highest mortality were hydramethylnon and fipronil. These data on the efficacy of commercially available contact and bait insecticides provide valuable information to manage this invasive pest.

Tunneling Performance Increases at Lower Temperatures for Solenopsis invicta (Buren) but not for Nylanderia fulva (Mayr).

Nylanderia fulva (Mayr), the tawny crazy ant, is an invasive pest established in Florida and several other Gulf Coast states. In their invasive ranges in the Southeastern USA, large N. fulva populations have reduced species abundance, even displacing another invasive ant, Solenopsis invicta (Buren). In North Florida, N. fulva populations survive winter temperatures that reach below freezing for extended periods. However, the shallow littoral debris used by N. fulva for nest construction offers little insulation to brood and reproductives when exposed to freezing temperatures. Field populations of N. fulva in North Florida were observed tunneling below ground, a previously undescribed behavior. Other invasive ants exhibit similar subterranean tunneling behavior as a means of thermoregulation. To test the hypothesis that N. fulva has the capacity to construct subterranean tunnels across a range of ecologically relevant temperatures, tunneling performance for N. fulva and S. invicta, another invasive ant that tunnels extensively, were compared at four temperatures (15.0, 18.0, 20.0, and 22.0 °C). Overall, N. fulva tunneled significantly less than S. invicta. Nylanderia fulva tunneled furthest at warmer temperatures whereas S. invicta tunneled furthest at cooler temperatures. However, N. fulva constructed subterranean tunnels at all temperatures evaluated. These data support the hypothesis that N. fulva is capable of tunneling in temperatures as low as 15.0 °C, confirming that this ant can also perform a behavior that is used by other ants for cold avoidance.