Nepetalactones from essential oil of Nepeta cataria represent a stable fly feeding and oviposition repellent.

The stable fly, Stomoxys calcitrans (L.) (Diptera: Muscidae), is one of the most serious pests to livestock. It feeds mainly on cattle and causes significant economic losses in the cattle industry. Standard stable fly control involving insecticides and sanitation is usually costly and often has limited effectiveness. As we continue to evaluate and develop safer fly control strategies, the present study reports on the effectiveness of catnip (Nepeta cataria L.) oil and its constituent compounds, nepetalactones, as stable fly repellents. The essential oil of catnip reduced the feeding of stable flies by >96% in an in vitro bioassay system, compared with other sesquiterpene-rich plant oils (e.g. amyris and sandalwood). Catnip oil demonstrated strong repellency against stable flies relative to other chemicals for repelling biting insects, including isolongifolenone, 2-methylpiperidinyl-3-cyclohexen-1-carboxamide and (1S,2'S)-2-methylpiperidinyl-3-cyclohexen-1-carboxamide. The repellency against stable flies of the most commonly used mosquito repellent, DEET, was relatively low. In field trials, two formulations of catnip oil provided >95% protection and were effective for up to 6 h when tested on cattle. Catnip oil also acted as a strong oviposition repellent and reduced gravid stable fly oviposition by 98%.

Determination by HPLC fluorescence analysis of the natural enantiomers of sex pheromones in the New World screwworm fly, Cochliomyia hominivorax.

Bioassays of six racemic synthesized candidate sex pheromone compounds against male New World screwworm Cochliomyia hominivorax (Coquerel) flies showed that the most potent bioactivity was found with 6-acetoxy-19-methylnonacosane and 7-acetoxy-15-methylnonacosane compared with four other isomeric acetoxy nonacosanes and a larger aliphatic ketone. As all these methyl-branched compounds have two asymmetric carbons and four possible enantiomers, characterization of the natural enantiomers was essential. All four enantiomers for the two most bioactive isomers of the natural sex pheromone were synthesized for bioassay. Hydrolysis and derivatization of these enantiomers with different fluorescent reagents was followed by column-switched high-performance liquid chromatography. The use of two linked, reversed-phase columns of different polarity held at sub-ambient temperatures allowed good separation of each enantiomer. This analysis applied to natural material was successful, as (6R,19R)-6-acetoxy-19-methylnonanocosane, and (7R,15R)- and (7R,15S)-7-acetoxy-15-methylnonanocosane were detected in extracts of recently colonized female flies.

Permeabilization of Cochliomyia hominivorax (Diptera: Calliphoridae) embryos.

Embryos of the primary screwworm, Cochliomyia hominivorax (Coquerel), were successfully permeabilized for use in subsequent cryopreservation studies. Mortality was greater for eggs incubated for < 5 h before treatment. The mean survival of embryos to first instars was 55.7, 61.1, and 62.6% when the embryos were incubated for 5, 5.5, and 6 h before treatment, respectively. The survival to the pupal and adult stages was low. An improved media for culturing the embryos during and immediately after treatment needs to be devised and the procedure for rearing the larval stages also needs to be altered to improve survival for emerging adults.

Identification of screwworms, Cochliomyia hominivorax (Coquerel) (Diptera: Calliphoridae), with a monoclonal antibody-based enzyme-linked immunosorbent assay (MAb-ELISA).

Myiasis caused by screwworms, Cochliomyia hominivorax (Coquerel), is devastating to warm-blooded animals and economically important to livestock producers. It is difficult to distinguish these pests, immature screwworms, from immatures of other non-pest fly species that often occur in animal wounds; it would be helpful to have tools available that do not rely on morphological characteristics. We developed two monoclonal antibodies (MAbs), highly specific for the screwworm, and used them in an enzyme-linked immunosorbent assay (MAb-ELISA), that differentiated screwworm eggs, larvae, pupae, and adults from those of the closely related secondary screwworm, C. macellaria (Fabricius) as well as Phormia regina (Meigen), Phaenicia sericata (Meigen), Calliphora vicina Robineau-Desvoidy, and Chrysomya rufifacies (Macquart). In a blind study, the microplate MAb-ELISA, which took about 4h to complete, displayed high specificity (99%), sensitivity (92%) and overall accuracy (97%) in distinguishing all life stages of the screwworm. Electrophoresis results suggested that the two monoclonal antibodies recognized identical conformational epitopes present in all screwworm life stages. The screwworm eradication program, successful in eradicating this pest from the US, Mexico, most of Central America and Libya (after an accidental introduction), could benefit in future eradication, surveillance, and exclusion efforts by developing a reliable field identification kit based on MAb-ELISA that accurately and quickly distinguished cases of screwworm myiasis.

Overwintering of the stable fly (Diptera: Muscidae) in southeastern Nebraska.

Adult stable flies, Stomoxys calcitrans (L.), were monitored during three winters at two, four, and 13 locations with Alsynite fiberglass traps and by examination of the interiors of buildings. No stable flies were found inside buildings during the winter. Adult stable flies were consistently caught on Alsynite traps at one location during two winters and at two other locations during one winter. Distribution and physiological age of these flies indicate that they emerged from pupae that had developed at or near the location where they were captured. Potential breeding sites were examined. A few third instars were found in silage in midwinter, but silage should not be considered a major source for overwintering stable flies.

Scheduled sanitation to reduce stable fly (Diptera: Muscidae) populations in beef cattle feedlots.

Sanitation has been long recommended as a means of reducing stable fly, Stomoxys calcitrans (L.), populations at cattle feedlots, but there is little published research to support this recommendation. In each of the 2 yr of this study, 4 feedlots received complete sanitation and 4 feedlots received no cleaning. The objective was to have the initial cleaning done before 1 June and then to reclean as needed every 2 wk thereafter. The feedlots that were cleaned had significantly fewer flies than the uncleaned feedlots, with 50.9% fewer stable flies during the 1st yr and 36.2% fewer flies in the 2nd yr. These reductions were realized even though initial cleaning was not done by 1 June in either year: bad weather delayed completion until 20 June the 1st yr and 29 June the 2nd yr. Sanitation at cattle feedlots significantly reduces stable fly populations and sanitation may have been more effective if initial cleaning was done by 1 June.

Effects of stable flies (Diptera: Muscidae) on weight gains of grazing yearling cattle.

Differences in weight gains caused by stable flies, Stomoxys calcitrans (L.), on grazing yearling steer/calves averaged 0.2 kg per steer in a 3-yr study on canyon range pastures in West Central Nebraska, Stable fly numbers averaged 0.85 per front leg on treated calves and 3.64 per front leg on control calves. In 2 of the 3 yr after the grazing trials were completed, the calves were placed in a feedlot and fed a finishing ration. Compensatory gain did not occur in the feedlot after the stable fly stress was removed.

Dispersal of stable flies (Diptera: Muscidae) from larval development sites in a Nebraska landscape.

Seven mark-recapture studies were conducted over 3 yr to assess dispersal of newly emerging adult stable flies, Stomoxys calcitrans L., from larval development sites in a mixed agricultural environment in northeastern Nebraska. Infested hay debris piles were marked by dusting their surfaces with fluorescent pigments, adults were captured with surrounding grids of Alsynite sticky traps, and specimens were dissected to determine feeding histories and reproductive age. Distances and directions of 3,889 marked specimens indicated males and females dispersed equally and in all directions. Midguts of males and females were equally likely to contain blood-meal remnants. Percentage with blood remnants and percentage of females with yolk increased with distance from mark origin, indicating survival and spread were positively associated with host finding success. A time-integrated diffusion model fit to results from the seven studies indicated 50% of stable fly adults had dispersed beyond 1.6 km of their natal site, but only 5% had dispersed beyond 5.1 km. These results indicate that stable fly adults on cattle in a given area are most likely to have originated from larval development sites within an ≈ 5 km radius of the subject cattle.

Candidate sex pheromones of the New World screwworm Cochliomyia hominivorax.

Five novel homologous acetate derivatives of long-chain secondary alcohols and a related ketone were tested for their efficacy as contact mating stimulants for Cochliomyia hominivorax Coquerel (Diptera: Calliphoridae). Full copulatory behaviour at a high percentage was found in tests with racemic 6-acetoxy-19-methylnonacosane at 2.5-20 microg using fertile males from three strains. Males of two strains responded nearly as well to 7-acetoxy-15-methylnonacosane, but an older strain first colonized in 1992 did not respond to this compound. Few or no copulatory responses were obtained to the other secondary alcohol acetates and a related ketone. These two acetate derivatives are the first sex pheromones identified in a calliphorid fly. The threshold of response was also tested, but could not be pinpointed.

piggyBac transformation of the New World screwworm, Cochliomyia hominivorax, produces multiple distinct mutant strains.

Sterile insect technique (SIT) programs are designed to eradicate pest species by releasing mass-reared, sterile insects into an infested area. The first major implementation of SIT was the New World Screwworm Eradication Program, which successfully eliminated the New World screwworm (NWS), Cochliomyia hominivorax (Coquerel) (Diptera: Calliphoridae), from the Continental US, Mexico and much of Central America. Ionizing radiation is currently used for sterilization, but transgenic insect techniques could replace this method, providing a safer, more cost-effective alternative. Genetic transformation methods have been demonstrated in NWS, and verified by Southern blot hybridization, PCR and sequencing of element insertion junctions. A lethal insertional mutation and enhancer detection-like phenotypic expression variations are presented and discussed. In addition to supporting the eradication efforts, transformation methods offer potential means to identify genes and examine gene function in NWS.