Prevalence and distribution of pathogen infection and permethrin resistance in tropical and temperate populations of Rhipicephalus sanguineus s.l. collected worldwide.

The brown dog tick, Rhipicephalus sanguineus sensu lato (s.l.) Latreille (Acari: Ixodidae), is a peridomestic, cosmopolitan parasite of dogs known to vector numerous pathogens of veterinary and medical importance. Recent phylogenetic analyses separate this tick into temperate and tropical lineages. Populations of Rh. sanguineus s.l. have been reported to exhibit sodium channel target site insensitivity to permethrin and etofenprox, which is likely due to the prolonged use of pyrethroids against many pests in and around the home. In this study, populations collected in the Caribbean, Africa, Asia, Europe and North America, were tested to identify the distribution of a known resistance mechanism, pathogen-vector interactions and phylogeny in relation to latitude. Using molecular assays, populations from 29 distinct locations were simultaneously geographically typed and screened for bacterial infection by Rickettsia, Ehrlichia, Babesia and Hepatozoon species, and for the presence of a sodium channel single nucleotide polymorphism known to confer permethrin resistance. Implications of these results on Rh. sanguineus s.l. management in association with geographical distribution will be discussed.

Auto-dissemination of commercially available fungal pathogens in a laboratory assay for management of the brown dog tick Rhipicephalus sanguineus.

Rhipicephalus sanguineus is a canine tick that infests dogs throughout the world and is frequently found in homes and dog kennels. Management of this tick species is complicated by the presence of resistance to commonly utilized acaricides. Fungal formulations could provide a valuable alternative tool for management and are especially relevant indoors where detrimental environmental effects on fungal spores are of less concern. Two commercially available fungal formulations, one containing Metarhizium anisopliae and the other containing Beauveria bassiana, were compared for time to death and sporulation in nymphal ticks exposed for 60 min in treated filter paper packets. Beauveria bassiana exposure killed ticks faster than M. anisopliae exposure and B. bassiana was more likely to sporulate on tick cadavers than M. anisopliae. To determine whether infected ticks could disseminate fungus to their conspecifics, ticks were marked and treated with fungus before being placed with untreated ticks. Fungus was successfully transmitted from treated to untreated ticks. Mortality of ticks exposed to B. bassiana-exposed conspecifics occurred sooner than for those exposed to M. anisopliae-exposed conspecifics, indicating faster dissemination in the former. Therefore, although both formulations resulted in decreased longevity of ticks compared with the controls, the B. bassiana formulation holds the most promise for direct or indirect application with respect to brown dog tick management.

Permethrin and malathion LD90 values for Culex quinquefasciatus vary with topical application site.

Prior research in multiple insect species has demonstrated that insecticide-induced mortality varies according to the body region exposed on the insect. This variation has been demonstrated in Culex quinquefasciatus Say (Diptera: Culicidae), but has not been quantified using dose-response curves. Applications of technical permethrin or malathion to one of three body regions on Cx. quinquefasciatus resulted in dose-response curves that were not equivalent to one another. The generated LD90 values and curves for each body region were compared with previously reported LD values for analogous sites in several mosquito species, specifically the mesothorax. Based on the present results, the permethrin and malathion LD50 and LD90 concentrations required for droplets impinging on the abdomen and mesothorax of Cx. quinquefasciatus when applied through ground-based spray systems utilized by mosquito control programmes were calculated.

Effects of four commercial fungal formulations on mortality and sporulation in house flies (Musca domestica) and stable flies (Stomoxys calcitrans).

The house fly Musca domestica L. (Diptera: Muscidae) and stable fly Stomoxys calcitrans (L.) (Diptera: Muscidae) are major pests of livestock. Biological control is an important tool in an integrated control framework. Increased mortality in filth flies has been documented with entomopathogenic fungi, several strains of which are commercially available. Three strains of Beauveria bassiana (Balsamo-Crivelli) Vuillemin (Hypocreales: Cordycipitaceae) and one strain of Metarhizium brunneum (Petch) (Hypocreales: Clavicipitaceae) were tested in commercial formulations for pathogenicity against house flies and stable flies. There was a significant increase in mortality of house flies with three of the formulations, BotaniGard® ES, Mycotrol® O, and Met52® EC, during days 4-9 in comparison with balEnce™ and the control. In stable flies, mortality rates were highest with Met52® EC, followed by Mycotrol® O, BotaniGard® ES and, finally, balEnce™. There was a significant fungal effect on sporulation in both house flies and stable flies. Product formulation, species differences and fungal strains may be responsible for some of the differences observed. Future testing in field situations is necessary. These commercial biopesticides may represent important tools in integrated fly management programmes.

Determination of metabolic resistance mechanisms in pyrethroid-resistant and fipronil-tolerant brown dog ticks.

Rhipicephalus sanguineus (Latreille) (Ixodida: Ixodidae) is a three-host dog tick found worldwide that is able to complete its' entire lifecycle indoors. Options for the management of R. sanguineus are limited and its' control relies largely on only a few acaricidal active ingredients. Previous studies have confirmed permethrin resistance and fipronil tolerance in R. sanguineus populations, commonly conferred by metabolic detoxification or target site mutations. Herein, five strains of permethrin-resistant and three strains of fipronil-tolerant ticks were evaluated for metabolic resistance using synergists to block metabolic enzymes. Synergist studies were completed with triphenyl phosphate (TPP) for esterase inhibition, piperonyl butoxide (PBO) for cytochrome P450 inhibition, and diethyl maleate (DEM) for glutathione-S-transferase inhibition. Additionally, increased esterase activity was confirmed using gel electrophoresis. The most important metabolic detoxification mechanism in permethrin-resistant ticks was increased esterase activity, followed by increased cytochrome P450 activity. The inhibition of metabolic enzymes did not have a marked impact on fipronil-tolerant tick strains.

Does behaviour play a role in house fly resistance to imidacloprid-containing baits?

The objective of this research was to examine the role and type of behavioural mechanisms that function in house fly, Musca domestica L. (Diptera: Muscidae), resistance to an imidacloprid-containing commercial fly bait, QuickBayt(®) , using an insecticide-susceptible and an imidacloprid-resistant strain. Mortality and feeding behaviour were observed through choice bioassays of three post-imidacloprid selected house fly generations to determine whether flies would consume the bait in the presence of an alternative food source. Mortality rates in choice containers progressively decreased in post-selection flies as QuickBayt(®) no-choice selections proceeded. There were no differences between the proportions of flies observed contacting QuickBayt(®) and sugar, respectively, a finding that eliminates repellency as a mechanism of stimulus-dependent behavioural resistance. However, differences in QuickBayt(®) consumption and subsequent mortality between choice and no-choice containers provided strong support for the evolution of consumption irritancy- or taste aversion-related behavioural resistance. The results of this study support the responsible rotation of insecticide bait formulations for house fly control.

Comparison of ectoparasite communities of sylvatic and urban wild mesomammals and unowned community cats in north-central Florida.

The adaptation of wildlife species to urban environments can drive changes in the ecology of ectoparasites and vector-borne disease. To better understand ectoparasite dynamics in an urban environment, we investigated the ectoparasite communities of 183 sylvatic and urban opossums and raccoons captured across four seasons at a rural research station and within the city of Gainesville, FL, and of 115 community cats from the Gainesville, FL area. Amblyomma americanum (L.) (Acari: Ixodidae), Dermacentor variabilis (Say), and Ixodes texanus Banks were collected from raccoons, A. americanum, D. variabilis, and Ixodes scapularis Say from opossums, and A. americanum from cats. Few ticks were collected from urban animals, although species richness of ectoparasites was similar between urban and sylvatic habitats. Ctenocephalides felis (Bouché) (Siphonaptera: Pulicidae) was collected from all sampled host species, but was particularly abundant on opossums. Additionally, Orchopeas howardi (Baker) (Siphonaptera: Ceratophyllidae) was collected from raccoons, and O. howardi and Xenopsylla cheopis (Rothschild) (Siphonaptera: Pulicidae) from opossums. Only raccoons were infested with raccoon lice, and only cats were infested with cat lice. Primarily opossums were infested with mites. Ectoparasite community composition varied by habitat, host species, and season; seasonal variation in ectoparasite communities differed between the sylvatic and urban habitats. While urban mesomammals did not appear to play an important role in supporting tick populations in an urban habitat, urban opossums appear to serve as an alternate host for large numbers of cat fleas, which may be an important consideration for treatment and control efforts against ectoparasites of companion animals.

Determination of the Discriminating Concentration Towards Permethrin for Surveying Resistance in Amblyomma americanum.

Amblyomma americanum Linnaeus (Ixodida: Ixodidae) is ubiquitously present throughout the southeastern United States and is capable of vectoring several pathogens. White-tailed deer are the main host for adult A. americanum. However, this tick species is a generalist that will feed on most vertebrates, including humans, deer, livestock, and pets. Management of this species can be challenging due to a lack of cost-effective strategies. Acaricides are often utilized, however, this may lead to pesticide resistance. The Food and Agriculture Organization of the United Nations (FAO) larval packet test (LPT) was performed on susceptible A. americanum to determine the lethal concentration (LC) and discriminating concentration (DC) values for permethrin. The FAO LPT was used at these pre-established values to compare levels of resistance in ticks collected from a captive deer farm and wild areas representing high and low permethrin exposure settings, respectively. Resistance ratios (RR) calculated from the LC values for the ticks collected from farmed and wild deer ranged between 1 and 2. A. americanum collected from farmed and wild deer were not found to be resistant, however, some samples had slightly elevated RRs as compared to the susceptible laboratory strain, which may suggest tolerance development. Although the A. americanum sampled in this study were not resistant to permethrin, the DC calculated in this study will allow for rapid evaluation of resistance in a permethrin resistance monitoring program such that alternate management strategies can be adopted if resistance is detected.

Infusion-baited ovitraps to survey ovipositional height preferences of container-inhabiting mosquitoes in two Florida habitats.

To ascertain mosquito species-specific oviposition height preferences, a study was conducted evaluating the response of field populations of container-inhabiting mosquitoes to water, oak (Quercus spp.),or oak-pine (Pinus spp.) infusion-baited ovitraps in four suburban and four sylvatic habitats in north central Florida. In total, 48 ovitraps, 24 suspended at each height of 1 or 6 m (near the ground or tree canopy, respectively), were monitored weekly for 5 mo. Throughout our study, we collected 13,276 mosquito eggs, representing five species from four genera, the most common being Aedes triseriatus (Say), Aedes albopictus Skuse, and Orthopodomyia signifera (Coquillett). Significantly more Ae. triseriatus and Ae. albopictus eggs were oviposited in containers with oak and oak-pine infusions compared with water alone. Significantly more Ae. albopictus eggs were recovered from traps at 1 m in suburban habitats, whereas more Ae. triseriatus eggs were recovered at 6 m in sylvatic habitats.

Host-seeking height preferences of Aedes albopictus (Diptera: Culicidae) in north central Florida suburban and sylvatic locales.

The response of Aedes albopictus to the BG-Sentinel, Omni-directional-Fay-Prince, and Mosquito MagnetX traps was evaluated in four suburban and four sylvatic habitats in north-central Florida to ascertain potential height preference of this species. These traps, which are primarily designed to attract diurnal mosquitoes, were set at 1 and 6 m and were evaluated during 40 trapping periods over 4 mo. We collected 45,640 mosquitoes, representing 26 species from 10 genera, the most common being Aedes albopictus, Ae. vexans, Coquilletidia perturbans, Culex nigripalpus, Aedes infirmatus, Ae. triseriatus, and Psorophora ferox. Although significantly more Ae. albopictus were captured at 1 m above ground than at 6 m, fewer were captured in sylvatic habitats than suburban habitats. Although not statistically different, the BG-Sentinel caught more Ae. albopictus compared with the other two traps regardless of locale. These results suggest that, although Ae. albopictus was captured as high as 6 m, the majority seek hosts at or below 1-m heights. This further supports prior research that, although Ae. albopictus has been shown to disseminate West Nile virus, it has not been implicated as a major vector for the virus, which is likely because of its propensity to feed on ground-dwelling hosts. The study also shows how trap type, trap heights, and environments influence sampling estimates when determining species abundance.

Evaluation of two commercial traps for the collection of Culicoides (Diptera: Ceratopogonidae).

Two types of commercial propane-powered traps, mosquito magnet (MM) (American Biophysics Corporation) MM-Freedom (Freedom) and MM-Liberty Plus (Liberty Plus), were evaluated for the collection of Culicoides. Trap preference and seasonal characteristics for the 3 major species, Culicoides furens, Culicoides barbosai, and Culicoides mississippiensis, were recorded from July 7, 2005, to July 24, 2006. Over 35 million Culicoides were captured during our study. When species were evaluated separately, analysis of overall mean trap collections yielded 5 months (February, March, June, September, and October) with significant trap effects. The Freedom trap captured more C. furens in June and October; the Liberty Plus trap captured more C. mississippiensis in February, March, and April, and more C. barbosai in September. The high numbers of Culicoides captured during our study suggest that the number of host-seeking Culicoides could potentially be reduced by continuous trapping during times when they are prevalent. Results of these investigations will be used to guide future control efforts.

Mosquitoes (Diptera: Culicidae) Collected From Residential Yards and Dog Kennels in Florida Using Two Aspirators, a Sweep Net, or a CDC Trap.

Mosquito surveillance typically uses Centers for Disease Control and Prevention (CDC) mosquito light traps baited with CO2. From January 2013 to March 2015, we sampled seven field sites using three active mosquito-trapping techniques (two different aspirators and a sweep net) and the stationary CO2-baited CDC mosquito light trap to determine mosquito capture efficacy for each technique. Sampling occurred in four suburban backyards and three dog kennel facilities near Gainesville, FL, USA; species collection and relative abundance were measured. A total of 32 species and 70,090 individual mosquitoes were collected, including a new record for Alachua County, Florida, Aedes hendersoni (Cockerell). The dominant (>5% of total capture) mosquito species collected during the study included Aedes atlanticus (Dyar and Knab), Aedes infirmatus (Dyar and Knab), Anopheles crucians Wiedemann, Culiseta melanura (Coquillett), Culex erraticus (Dyar and Knab), Culex nigripalpus Theobald, and Uranotaenia sapphirina (Osten Sacken). The CDC trap captured the most species (29), followed by large aspirator (28), small aspirator (26), and the sweep net (23). All dominant species were captured with each sampling technique. Excluding Wyeomyia mitchellii (Theobald), all subdominant species (1-5% of total capture) were collected with each sampling technique. Future sampling should consider the utility (e.g., large numbers are readily collected) and limitations (e.g., personnel requirements) of aspirator collections when designing field-based mosquito sampling projects, especially those in residential areas or those focused upon species captured.

Development of Spalangia cameroni and Muscidifurax raptor (Hymenoptera: Pteromalidae) on live house fly (Diptera: Muscidae) pupae and pupae killed by heat shock, irradiation, and cold.

The objective of this study was to evaluate the suitability of killed house fly (Musca domestica L) pupae for production of two economically important pupal parasitoids. Two-day-old fly pupae were subjected to heat shock treatments of varying temperatures and durations in an oven at >or=70% RH; exposure to temperatures of 55 degrees C or higher for 15 min or longer resulted in 100% mortality. Exposure to 50 degrees C resulted in 40 and 91% mortality at 15 and 60 min, respectively. All (100%) pupae placed in a -80 degrees C freezer were killed after 10-min exposure; exposure times of <5 min resulted in <21% mortality. Progeny production of Spalangia cameroni Perkins and Muscidifurax raptor Girault and Sanders (Hymeoptera: Pteromalidae) from pupae killed by heat shock or 50 kR of gamma radiation was not significantly different from production on live hosts on the day when pupae were killed. Freeze-killed pupae produced 16% fewer S. cameroni than live pupae and an equivalent amount of M. raptor progeny on the day when pupae were killed. When killed pupae were stored in freezer bags at 4 degrees C for 4 mo, heat-killed, irradiated, and freeze-killed pupae remained as effective for production of M. raptor as live pupae. Production of S. cameroni on heat-killed and irradiated pupae was equal to parasitoid production on live pupae for up to 2 mo of storage, after which production on killed pupae declined to 63% of that observed with live pupae. Production of S. cameroni on freeze-killed pupae was 73-78% of production using live pupae during weeks 2-8 of storage and declined to 41 and 28% after 3 and 4 mo, respectively. Killing pupae by heat shock provides a simple and low-cost method for stockpiling high-quality hosts for mass-rearing both of these filth fly biological control agents.

Mutation in the Sodium Channel Gene Corresponds With Phenotypic Resistance of Rhipicephalus sanguineus sensu lato (Acari: Ixodidae) to Pyrethroids.

The brown dog tick, Rhipicephalus sanguineus sensu lato (Latreille), is a cosmopolitan ectoparasite and vector of pathogens that kill humans and animals. Pyrethroids represent a class of synthetic acaricides that have been used intensely to try to control the brown dog tick and mitigate the risk of tick-borne disease transmission. However, acaricide resistance is an emerging problem in the management of the brown dog tick. Understanding the mechanism of resistance to acaricides, including pyrethroids, is important to adapt brown dog tick control strategies. The main objective of this study was to determine if target-site mutations associated with pyrethroid resistance in other pests could be associated with phenotypic resistance detected in a brown dog tick population from Florida. We amplified segment 6 of the domain III of the voltage-sensitive sodium channel protein, using cDNAs synthesized from pyrethroid-susceptible and pyrethroid-resistant tick strains. A single nucleotide point mutation (SNP) identified in a highly conserved region of domain III S6 in the resistant ticks resulted in an amino acid change from phenylalanine to leucine. This mutation is characteristic of resistance phenotypes in other tick species, and is the first report of this mutation in R. sanguineus. Molecular assays based on this knowledge could be developed to diagnose the risk for pyrethroid resistance, and to inform decisions on integrated brown dog tick management practices.

Seasonal variation in Carcinops pumilio (Coleoptera: Histeridae) dispersal and potential for suppression of dispersal behavior.

Seasonal dispersal of Carcinops pumilio (Erichson) was evaluated using two trapping methods-a black-light pitfall trap and a mesh-bottomed trap placed on poultry manure. The black-light trap collected larger numbers than the mesh-bottomed trap from March through June. The mesh-bottomed trap gathered larger numbers of beetles from June through August and numbers were less variable throughout the year. Often, when very low numbers of beetles were recovered from manure cores, large numbers of beetles could be collected with the black-light trap suggesting that beetle density may not be an important factor in dispersal behavior. The greatest dispersal in the dispersal arenas (approximately 90%) occurred using beetles collected by both trap types in June 2000. Beginning in March and ending in August, a cyclic rise and then fall pattern in both laboratory dispersal and beetle collections was observed. Trap collection patterns were similar in both years of the study. In January and March, we were unable to prevent dispersal behavior of beetles captured in black-light traps. However, in May, after beetles had been in a dispersal phase for several months, we were able to suppress dispersal. In contrast, dispersal behavior among beetles captured with the mesh-bottomed trap did not change following the photoperiod-altered exposure.

Larval production from field-collected Carcinops pumilio (Coleoptera: Histeridae) following three starvation periods.

Carcinops pumilio (Erichson) were collected from high-rise, caged-layer poultry facilities using two trapping methods, a blacklight pitfall trap and a mesh-bottomed trap placed on poultry manure. Starvation for 14 d significantly reduced larval production during the first 3-d oviposition period regardless of trapping method. Beetles collected with blacklight traps and subsequently starved for 14 d had higher larval production in the third through fifth oviposition periods than those fed daily, indicating that lack of nutrition was a limiting factor in C. pumilio larval production. No differences were observed in larval production, after the first oviposition period, between the 14-d starved and daily fed groups collected with the mesh-bottom trap. In all blacklight-captured treatments, larval production was lowest during the first oviposition period with the largest differences found among the three starved treatments. Larval production in the 14-d starved treatment increased significantly during the later oviposition periods in mesh-bottom trap studies. Within the fed treatment, larval production was consistently greater among beetles collected with the mesh-bottom trap than among beetles collected with blacklight traps.

Prey- and density-mediated dispersal in Carcinops pumilio (Coleoptera: Histeridae), a predator of house fly (Diptera: Muscidae) eggs and larvae.

Carcinops pumilio (Erichson) were collected using two trapping methods: a black light pitfall trap and a mesh-bottomed trap placed on poultry manure. C. pumilio collected with black lights and subsequently starved had a significantly higher dispersal rate during days 1-3 than fully fed groups. When densities of < 500 Caloglyphus berlesei (Michael) (Acarina: Acaridae) per 50 C. pumilio were provided, mite availability had a significant effect on dispersal of beetles captured with both black light traps and the mesh-bottomed trap during the day 1-3 period. Our results indicate that the availability of acceptable food sources can delay and possibly prevent dispersal by C. pumilio. Black light-captured beetles appeared to be in a state of dispersal when captured. A subset of dispersing beetles was present in groups captured with the mesh-bottomed trap.

Efficacy of chlorfenapyr (AC 303630) experimental pour-on and CyLence formulations against naturally acquired louse infestations on cattle in New York.

The four chlorfenapyr formulations examined provided 100% control of both the nymphal and adult stages of naturally acquired Bovicola bovis (L.) on cattle up to 35 days after application. Treatment with 6mg chlorfenapyr per kg BW in a 0.12ml per kg BW formulation was as effective as treatment with CyLence (cyfluthrin) in controlling naturally acquired Solenopotes capillatus (Enderlein) on cattle for 35 days. Percent reduction was never greater than 90% with any chlorfenapyr application against Linognathus vituli (L.). However, percent reduction was greater than 90% with CyLence from day 21 through 35. No adverse effects were noted on cattle from any of the chlorfenapyr dosages used.

Monitoring insecticide resistance in house flies (Diptera: Muscidae) from New York dairies.

House flies were collected from dairies across New York state and the levels of resistance to seven insecticides were measured using standard laboratory assays with three to five diagnostic concentrations. The highest levels of resistance were found for tetrachlorvinphos, permethrin and cyfluthrin. Although levels of resistance to methomyl were somewhat lower, they were among the highest ever reported for field-collected house flies. Resistance to pyrethrins was limited primarily to the lowest diagnostic concentration. House flies were susceptible to fipronil at all dairies, suggesting that this material would be highly effective for fly control. The levels of resistance were similar at all the dairies, irrespective of their insecticide use, suggesting substantial movement of flies between facilities. Relative to resistance levels found at New York dairies in 1987, resistance levels had increased for permethrin, were unchanged for tetrachlorvinphos and had decreased for dimethoate. To identify a single diagnostic concentration that could be used in the laboratory assays to assess accurately resistance levels in future studies, we carried out a 'simulated' field bioassay using formulated materials. A diagnostic concentration for each insecticide is proposed on the basis of a comparison of our bioassays.

Impact of exposure length and pupal source on Muscidifurax raptorellus and Nasonia vitripennis (Hymenoptera: Pteromalidae) parasitism in a New York poultry facility.

Commercially obtained Nasonia vitripennis Walker and Muscidifurax raptorellus Kogan & Legner were released weekly for 12 wk into a high-rise, caged-layer poultry house. After the release period, parasitoids were sampled using sentinel house fly (Musca domestica L.) pupae that were either laboratory-reared or field-collected as larvae and exposed for 2, 4, 7, and 14 d. Parasitoid-induced mortality was observed in 31% of laboratory colony pupae and in 26% of field-collected pupae, whereas successful parasitism rates of 48 and 51% were observed from these pupal sources, respectively. Parasitism was primarily by M. raptorellus (88%), and Muscidifurax raptor Girault & Sanders (11%), while N. vitripennis accounted for <1%. Percent female progeny ranged from 43%, in M. raptorellus to 76% in N. vitripennis. Parasitoid emergence from 2-d exposed sentinel pupae was the lowest of all treatments. Parasitoid emergence from 7-d exposed sentinel pupae was the highest of all treatments. We found no differences between pupal source, suggesting that when sampling for M. raptor, M. raptorellus, and N. vitripennis, in poultry facilities, pupal source is not a confounding factor.