International Program to Monitor Cat Flea Populations for Susceptibility to Imidacloprid.

An international team of scientists and veterinarians was assembled in 1999 to develop a monitoring program to determine the susceptibility of cat fleas, Ctenocephalides felis felis (Bouché) (Siphonaptera: Pulicidae), to imidacloprid. Cat flea eggs were collected, shipped to laboratories, and tested for their susceptibility to imidacloprid. Over 3,000 C. felis populations were collected from 2002 to 2017 from 10 different countries. Of these, 66.3% were collected from cats and 33.7% from dogs. C. f. felis populations (n = 2,200) were bioassayed by exposing cat flea eggs and the emerging larvae to a Diagnostic Dose (DD) of 3 ppm imidacloprid in larval rearing medium. Flea eggs hatched and developed in the untreated controls in 1,837 of the isolates (83.5%) bioassayed. Flea isolates (n = 61) that had ≥5% survival at the DD of 3 ppm were retested with a second DD of 3 ppm. None of them had ≥5% survival to the second dose of 3 ppm. Of the 1,837 valid C. felis isolates tested, there has been no evidence of a decreased susceptibility to imidacloprid over the past 17 yr. The methods outlined in this article should provide an acceptable protocol for testing many of the new active ingredients that have been registered for cat flea control.

Insecticide Transfer Efficiency and Lethal Load in Argentine Ants.

Trophallaxis between individual worker ants and the toxicant load in dead and live Argentine ants (Linepithema humile) in colonies exposed to fipronil and hydramethylnon experimental baits were examined using accelerator mass spectrometry (AMS). About 50% of the content of the crop containing trace levels of 14C-sucrose, 14C-hydramethylnon, and 14C-fipronil was shared between single donor and recipient ants. Dead workers and queens contained significantly more hydramethylnon (122.7 and 22.4 amol/µg ant, respectively) than did live workers and queens (96.3 and 10.4 amol/µg ant, respectively). Dead workers had significantly more fipronil (420.3 amol/µg ant) than did live workers (208.5 amol/µg ant), but dead and live queens had equal fipronil levels (59.5 and 54.3 amol/µg ant, respectively). The distribution of fipronil differed within the bodies of dead and live queens; the highest amounts of fipronil were recovered in the thorax of dead queens whereas live queens had the highest levels in the head. Resurgence of polygynous ant colonies treated with hydramethylnon baits may be explained by queen survival resulting from sublethal doses due to a slowing of trophallaxis throughout the colony. Bait strategies and dose levels for controlling insect pests need to be based on the specific toxicant properties and trophic strategies for targeting the entire colony.

Susceptibility of cat fleas (Siphonaptera: Pulicidae) to fipronil and imidacloprid using adult and larval bioassays.

The monitoring of the susceptibility offleas to insecticides has typically been conducted by exposing adults on treated surfaces. Other methods such as topical applications of insecticides to adults and larval bioassays on treated rearing media have been developed. Unfortunately, baseline responses of susceptible strains of cat flea, Ctenocephalides felis (Bouchè), except for imidacloprid, have not been determined for all on-animal therapies and new classes of chemistry now being used. However, the relationship between adult and larval bioassays of fleas has not been previously investigated. The adult and larval bioassays of fipronil and imidacloprid were compared for both field-collected isolates and laboratory strains. Adult topical bioassays of fipronil and imidacloprid to laboratory strains and field-collected isolates demonstrated that LD50s of fipronil and imidacloprid ranged from 0.11 to 0.40 nanograms per flea and 0.02 to 0.18 nanograms per flea, respectively. Resistance ratios for fipronil and imidacloprid ranged from 0.11 to 2.21. Based on the larval bioassay published for imidacloprid, a larval bioassay was established for fipronil and reported in this article. The ranges of the LC50s of fipronil and imidacloprid in the larval rearing media were 0.07-0.16 and 0.11-0.21 ppm, respectively. Resistance ratios for adult and larval bioassays ranged from 0.11 to 2.2 and 0.58 to 1.75, respectively. Both adult and larval bioassays provided similar patterns for fipronil and imidacloprid. Although the adult bioassays permitted a more precise dosage applied, the larval bioassays allowed for testing isolates without the need to maintain on synthetic or natural hosts.

Large-scale monitoring of imidacloprid susceptibility in the cat flea, Ctenocephalides felis.

Although on-animal topical treatment with compounds such as imidacloprid has revolutionized the control of the cat flea, Ctenocephalides felis (Bouché) (Siphonaptera: Pulicidae), the development of insecticide resistance is a continuing threat. As part of a highly co-ordinated and unprecedented resistance monitoring programme for C. felis, 1437 flea isolates were collected by veterinary clinics in Australia, Germany, France, the U.K. and 29 states in the U.S.A. from 2002 to 2009. About 65% of the collections were made from June to October each year and 71% of the collections were from cats. Collections of flea eggs were sent to one of five different laboratories, where they were tested with a diagnostic dose of imidacloprid (3 p.p.m.) applied to larval flea-rearing medium. Of the 1437 collections received, 1064 contained adequate numbers of eggs for testing. Of these isolates, untreated eggs failed to hatch in 22.7% and were not considered valid bioassays. Survival rates >5% and development of adult fleas (a threshold for further testing) occurred in only 22 isolates. They were re-tested with the same diagnostic dose and none produced >5% adult emergence. Complete dose-response bioassays were performed on three of the isolates that had triggered a second test and produced slopes, intercepts and LC(50) values similar to those for existing susceptible laboratory strains. Results confirmed sustained susceptibility of C. felis to imidacloprid, despite its widespread use for over a decade.

Toxicity, repellency, and effects of acetamiprid on western subterranean termite (Isoptera: Rhinotermitidae).

The insecticidial and biological activity of the cyano-substituted neonicotinoid acetamiprid was determined against the western subterranean termite, Reticulitermes hesperus Banks (Isoptera: Rhinotermitidae). Acetamiprid was very active against termites in topical applications, with an LD50 = 0.02 ng per termite. Even though acetamiprid was extremely toxic in topical applications, deposits > or =50 ppm on sand were required to consistently provide >90% kill of termites within 7 d after a 1-h exposure. Termites were quickly affected by brief exposures to sand treated with 1 ppm acetamiprid and within 1 h, their locomotion was dramatically impaired. Acetamiprid was transferred from donors to recipients only when donors were held on deposits > or =50 ppm for 1 h. Deposits even as low as 1 ppm were repellent and termites failed to tunnel into treated sand, and there was no significant mortality. Exposure to acetamiprid impaired locomotion of termites as did other slow-acting neonicotinoids, such as imidacloprid. Acetamiprid was repellent at all concentrations tested, acting like type I pyrethroid treatments in soil. A new subcategory of type III soil termiticides is proposed that incorporates the sublethal and delayed effects observed in neonicotinoid insecticides, and repellency at certain concentrations.

Aggressive interactions between Solenopsis invicta and Linepithema humile (Hymenoptera: Formicidae) under laboratory conditions.

The Argentine ant, Linepithema humile (Mayr), and the red imported fire ant, Solenopsis invicta Buren, are natural agonists in their country of origin. Since the first report of L. humile in California in 1907 its range expanded statewide, displacing native ant species wherever it spread. Since the discovery of established populations of S. invicta in southern California in 1998, it has been restricted to discrete areas of southern California. However, as these discrete populations expand, they are encountering large populations of L. humile, which are effective competitors for available resources and are particularly aggressive in their encounters with other ant species such as S. invicta. Most Dolichoderine ants such as L. humile do not prefer to forage on baits made with defatted corn grit and soybean oil typically used in red imported fire ant control programs. Applications of these baits in areas where distributions of these species overlap might selectively affect populations of S. invicta and give L. humile a competitive advantage. Three laboratory experiments were conducted to determine the competitive outcomes between S. invicta pitted against L. humile: 1) agonistic behavior of workers in small arenas, 2) colony interactions with different population ratios, and 3) the effects of pyriproxyfen on the competitiveness of S. invicta against L. humile. Populations of S. invicta consisting of major workers killed more L. humile than did minors or a mixture of majors and minors. When paired against L. humile colonies consisting of 1,100 workers, colonies consisting of 38 S. invicta workers were easily defeated by L. humile. Colonies consisting of 450 S. invicta workers plugged their nest entrances, but they were ultimately defeated by L. humile after 13 d. The S. invicta colonies consisting of 1,100 workers took control of the bridge connecting the colonies, invaded the L. humile colony, killed the Argentine ant queens, and removed their brood. Pyriproxyfen-treated fire ant workers took significantly longer to chase the Argentine ants from a connecting bridge than did untreated fire ants. Thus, fire ant baits may have long-term effects on intercolonial aggression between S. invicta and L. humile, especially when Argentine ant populations are high in the summer.

Determining a diagnostic dose for imidacloprid susceptibility testing of field-collected isolates of cat fleas (Siphonaptera: Pulicidae).

The susceptibility of four laboratory strains of cat fleas, Ctenocephalides felis (Bouche), to imidacloprid was determined by three different laboratories, by using a standardized bioassay protocol. The probit lines generated by the different laboratories were very similar, with LC50 values ranging from 0.32 to 0.81 ppm. Based on these data, a diagnostic dose (DD) of 3 ppm imidacloprid in larval rearing media was provisionally identified for detecting shifts in tolerance, possibly as a consequence of incipient imidacloprid resistance. None of the larvae from the susceptible laboratory strains survived the DD. Eighteen field-collected isolates were evaluated for their susceptibility to imidacloprid and to validate a DD of 3 ppm. Probit lines from 18 field-collected isolates were very similar, with LC50 values ranging from 0.14 to 1.52 ppm. When exposed to the DD, between 3 and 10% of the exposed larvae emerged as adults from only three of the 18 isolates. All other field isolates gave 100% mortality at the DD. Under the criteria established (>5% survivorship at 3 ppm), two isolates would be established on mammalian hosts and more extensive tests conducted to exclude or confirm the presence of resistance. The DD of 3 ppm is robust enough to eliminate most of the susceptible isolates collected until today, yet low enough to identify possible isolates for further testing.

Molecular phylogeny of isolates of Ctenocephalides felis and related species based on analysis of ITS1, ITS2 and mitochondrial 16S rDNA sequences and random binding primers.

The phylogenetic relationships among 31 different flea isolates representing seven different species were studied by nucleotide sequence comparison of the internal transcribed spacer 1 (ITS1), internal transcribed spacer 2 (ITS2) and/or mitochondrial 16S ribosomal RNA gene (mt16S-rDNA) to examine the patterns of variation. Results show that all regions are useful in discriminating among flea species. In Ctenocephalides felis and Tunga penetrans, some differences in these gene regions occurred among different isolates within the same species. In the latter case, the differences are in the mt16S-rDNA region, with one isolate showing 48% divergence in nucleotide sequence. The taxonomic implications of this result are unclear at present. The gene regions revealed differences between C. felis isolates only after DNA sequencing the PCR products. Further differentiation among C. felis isolates was obtained using four different random binding primers (decamers) and primers for mammalian aldolase to amplify narrow differences in the genome. Using these primers we were able to discriminate between different C. felis isolates and determine that some of the genetic variation coincided with minor differences in response to the control agent imidacloprid. However, overall findings do not support the existence of subspecies of C. felis.

Efficacy and longevity of nitenpyram against adult cat fleas (Siphonaptera: Pulicidae).

Nitenpyram (Capstar) is a fast acting, orally administered flea treatment that is absorbed into the blood of the host animal and is readily available for uptake by feeding fleas. We examined the efficacy of a single dose of nitenpyram against adult cat fleas, Ctenocephalides felis (Bouché), over several days. We recorded adult flea mortality and flea egg production on treated and untreated cats. Nitenpyram provided 100% kill of all fleas on the host at the time of treatment and for up to 24 h after treatment. Between 24 and 48 h after treatment, there was a 98.6% reduction in adult flea numbers. From 48 to 72 h, there was a 5% reduction in adult fleas. There was a 97% reduction and 95.2% reduction in the number of flea eggs collected from treated versus untreated animals during the first 48 h and from 48 to 72 h, respectively. In addition, we quantified three distinct behavioral responses of infested adult cats treated with nitenpyram to determine the extent of any immediate, overt behavioral responses in treated animals. A significant increase in scratching, biting, licking, and twitching occurred for 5 h. The biting and licking continued for 7 h after treatment. Administration of nitenpyram provides an effective mechanism to eliminate adult fleas from hosts for up to 48 h after treatment.

Determining the foraging range and origin of resurgence after treatment of Argentine ant (Hymenoptera: Formicidae) in urban areas.

The foraging range and distribution of Argentine ant, Linepithema humile (Mayr), colonies in urban areas of southern California extended at least 61 m (200 feet) from feeding stations and structures. Ants were fed at 25% sucrose feeding stations containing 0.01% fluorescent brighter (FB28). Within 14 d, from 77-90% of the ants sampled next to the feeding stations were positive for FB28. The percentage of ants with FB28 declined gradually to approximately 55% 61 m away from the feeding station. The percentage of marked ants in the controls didn't change over the 4-wk-test period. There were approximately 290,000 ants visiting the monitored stations each night before treatments. The 0.0001% fipronil baits and 0.06% fipronil SC sprays provided significant reductions in at least 4 wk. The percentage of ants marked with FB28 decreased significantly in both bait and spray treatments over 4 wk suggesting that the resurgence of ants in treated areas were because of immigration from untreated areas. It is likely that much larger areas will need to be treated to control Argentine ants in urban settings, especially if baits are being used.

Development of a larval bioassay for susceptibility of cat fleas (Siphonaptera: Pulicidae) to imidacloprid.

Strategies for controlling cat fleas, Ctenocephalidesfelisfelis (Bouché), have undergone dramatic changes in the past 5 yr. With the advent of on-animal treatments with residual activity the potential for the development of insecticide resistance increases. A larval bioassay was developed to determine the baseline susceptibility of field-collected strains of cat fleas to imidacloprid. All four laboratory strains tested showed a similar level of susceptibility to imidacloprid. Advantages of this bioassay are that smaller numbers of fleas are required because flea eggs are collected for the test. Insect growth regulators and other novel insecticides can also be evaluated. Using a discriminating dose, the detection of reduced susceptibility in field strains can be determined with as few as 40 eggs.

An oral bioassay for the toxicity of hydramethylnon to individual workers and queens of Argentine ants, Linepithema humile.

We have developed an oral bioassay to determine the toxicity of hydramethylnon to individual workers and queens of the Argentine ant, Linepithema humile. We fed seven concentrations of hydramethylnon in suspension to individual workers or queens, determined the amount of hydramethylnon ingested and evaluated the individual ants for mortality 14 days later. At concentrations > or = 0.37 g liter-1, the amount of liquid the queens ingested decreased dramatically, indicating that Argentine ant queens may detect hydramethylnon. Significantly larger volumes of the two highest concentrations of the hydramethylnon suspension were ingested by the workers, compared to the lower concentrations, suggesting that hydramethylnon may act as a feeding stimulant for the workers. Worker mortality was higher than queen mortality at the highest concentrations tested. The highest worker mortality resulted when the ants ingested 1.03 micrograms of hydramethylnon per mg of ant tissue. At the highest concentration (1.0 g liter-1) tested, workers ingested almost 12 times as much active ingredient per mg of body weight as did queens, suggesting that, in order to increase mortality of queens, multiple feedings must occur.

Laboratory techniques for rearing the fleas (Siphonaptera: Ceratophyllidae and Pulicidae) of California ground squirrels (Rodentia: Sciuridae) using a novel nest box.

Three species of fleas, Oropsylla montana (Baker), Hoplopsyllus anomalus (Baker), and Echidnophaga gallinacea (Westwood), occur seasonally on the California ground squirrel, Spermophilus beecheyi (Richardson). Few studies have focused on the biology and ecology of these fleas despite their importance in the epidemiology of sylvatic plague. To best duplicate a natural parasite-host relationship in the laboratory, a novel nest box was developed that facilitated housing wild-caught S. beecheyi, was conducive to rearing fleas, and met modern standards for laboratory animal hygiene. This flexible system allowed adult fleas with different feeding strategies to be colonized successfully while providing sufficient flea eggs for both colony maintenance and biological research. The techniques described could be adapted to work with other species of rodents and their fleas.

Toxicity and repellency of borate-sucrose water baits to Argentine ants (Hymenoptera: Formicidae).

The oral toxicity of boron compounds to the Argentine ant, Linepithema humile (Mayr), was evaluated in laboratory tests. The ants were provided 25% sucrose water containing 0.5 and 1% boric acid, disodium octaborate tetrahydrate, and borax. Lethal times of these solutions were a function of the concentration of boron. In field tests, the ants showed no discrimination between disodium octaborate tetrahydrate and boric acid. There was a significant reduction in consumption of sucrose water with > 1% boric acid.

Oral toxicity of abamectin, boric acid, fipronil, and hydramethylnon to laboratory colonies of Argentine ants (Hymenoptera: Formicidae).

Comprehensive laboratory baiting studies with the 4 insecticides abamectin, boric acid, fipronil, and hydramethylnon were conducted against Argentine ant, Linepithema humile (Mayr). Serial dilutions of each insecticide prepared in a 25% sucrose solution were provided for 24 h or 14 d to groups of workers and queens. Sucrose-water baits containing 1 x 10(-5)% fipronil and 0.1% hydramethylnon provided complete mortality of L. humile workers baited for 24 h. Baits containing 1 x 10(-4) and 1 x 10(-5)% fipronil provided 100% kill of queens. In the continuous baiting, 0.5% boric acid, 1 x 10(-6)% fipronil, and 0.025% hydramethylnon provided 100% mortality of workers. The queens and workers provided baits containing 1 x 10(-5)% fipronil and 0.5% boric acid all died within 14 d. Low concentrations of fipronil and boric acid in sucrose may be promising candidates for Argentine ant control.

Review of insecticide resistance in cat fleas (Siphonaptera: Pulicidae).

Insecticide resistance often is blamed for failures of insecticides to control cat fleas, Ctenocephalides felis (Bouché). Yet the genetics and adaptive advantage of resistance traits remain unexamined. Lethal doses of insecticides that kill 50% of the population fluctuate 7-fold within a cat flea strain. Many reports of flea resistance may be attributable to variable mortality from effects of solvents, substrates, humidities, temperatures, colonization, and ages of fleas. Resistance ratios (ratios of lethal doses of a resistant to a susceptible strain) are < 690-fold in fleas; lower than many other arthropods. This, plus strain variability, hinders resistance detection. Relationships between resistance levels, control failures, and health threats are unclear. Insensitive acetylcholinesterase, knockdown recovery, glutathione transferase conjugation, and mixed function oxidase/cytochrome P450 are demonstrated resistance mechanisms in cat fleas. Ecological genetics of resistance in cat fleas probably involves flea transfer among hosts, host movements, refugia, founder effects, and mortality from abiotic factors. Understanding cat flea resistance requires population monitoring before, during, and after insecticide treatments using conventional and rapid molecular bioassays. Sustained insecticide release devices such as flea collars and long-lived insecticide residues for premises possibly contribute to the development of resistance. New systemic and topical insecticides, especially when given prophylactically, may act similarly. Eliminating insecticides prevents insecticide resistance but necessitates application of biorational tactics incorporating mechanical, environmental, and cultural controls. Using high temperatures, low humidities, host grooming and such tactics as decreasing doses, increasing action thresholds, rotating insecticides, and leaving spatial and temporal refugia may suppress cat flea resistance.

Effect of temperature on cat flea (Siphonaptera:Pulicidae) development and overwintering.

Cat fleas, Ctenocephalides felis (Bouché), were reared at different temperatures to examine further the effects on egg-to-adult development and emergence of adults from cocoons. Three different temperatures (26.7, 21.1, and 15.5 degrees C) were used to rear fleas. Emergence of adults from cocoons was monitored using 2 experimental designs-cocoons maintained in groups and cocoons individually isolated. At cooler temperatures, cocoons kept in groups had a higher incidence of late-emerging adults than did individual cocoons. At 15.5 degrees C, adults continued to emerge from grouped cocoons up to 80 d after all had emerged from individual cocoons. Developmental time of prepupae and pupae was determined by inhibiting cocoon formation of late 3rd instars. Male prepupae and pupae developed approximately 20% slower than did females. Comparisons between emergence of adults from cocoons and from naked pupae indicate that a large percentage of adults emerge from cocoons soon after molting. The possible association of developmental temperatures and emergence from cocoons with overwintering populations is discussed.

The biology, ecology, and management of the cat flea.

The cat flea, Ctenocephalides felis felis, is the most important ectoparasite of domestic cats and dogs worldwide. In addition to its annoyance to pets and humans, C. felis felis is responsible for flea bite allergy dermatitis and the transmission of dog tapeworm. The abiotic and biotic factors that affect the development of immature stages are reviewed with special emphasis given to those aspects directly affecting control. Factors influencing host selection and feeding by adults are summarized. Recent studies concerning mating and oviposition, especially as they impact the likelihood of survival by immatures, are discussed. There has been an increase in the number of reports of insecticide resistance in the past ten years. Greater attention has been placed on disrupting larval development in modern IPM programs. The immature stages of the cat flea are extremely susceptible to environmental factors such as temperature and relative humidity and insect growth regulators (IGRs). In recent years, the control of cat fleas has increasingly relied on the use of IGRs applied to the host or to the indoor environment. Finally, we discuss advances in pesticide chemistry that provide tools for better control of adult fleas on the host.

Activity of insecticides applied to turfgrass to control adult cat fleas (Siphonaptera:Pulicidae).

A bioassay was developed to determine the efficacy of insecticides against adult cat fleas, Ctenocephalides felis (Bouché), on turfgrass. Activity was assessed by exposing adult fleas to treated and untreated disks of turf for 24 h, 1 and 7 d after treatment. Filter paper, a representative nonwaxy, porous substrate was bioassayed in similar fashion 1 d after treatment to confirm insecticide activity. Results demonstrated the relative efficacy of insecticides applied to turf at different rates as well as differences among insecticides. Of 10 insecticides tested, chlorpyrifos wettable powder and emulsifiable concentrates consistently provided greatest kill of adult fleas. Although initially active, pyrethroid insecticides and diazinon generally showed significant declines in residual activity by day 7. Possible reasons for differences in activity are discussed.