A genomics-informed,SNP association study reveals FBLN1 and FABP4 as contributing to resistance to fleece rot in Australian Merino sheep

Background  

Fleece rot (FR) and body-strike of Merino sheep by the sheep blowfly Lucilia cuprina are major problems for the Australian wool industry, causing significant losses as a result of increased management costs coupled with reduced wool productivity and quality. In addition to direct effects on fleece quality, fleece rot is a major predisposing factor to blowfly strike on the body of sheep. In order to investigate the genetic drivers of resistance to fleece rot, we constructed a combined ovine-bovine cDNA microarray of almost 12,000 probes including 6,125 skin expressed sequence tags and 5,760 anonymous clones obtained from skin subtracted libraries derived from fleece rot resistant and susceptible animals. This microarray platform was used to profile the gene expression changes between skin samples of six resistant and six susceptible animals taken immediately before, during and after FR induction. Mixed-model equations were employed to normalize the data and 155 genes were found to be differentially expressed (DE). Ten DE genes were selected for validation using real-time PCR on independent skin samples. The genomic regions of a further 5 DE genes were surveyed to identify single nucleotide polymorphisms (SNP) that were genotyped across three populations for their associations with fleece rot resistance.     

Habitat preferences and carcase colonization by sheep blowflies in the northern North Island of New Zealand

Abstract. In urban Auckland, from September 1991 to May 1992, only six specimens of the Australian sheep blowfly, Lucilia cuprina, were collected in farmed parkland and in garden habitats. The common green blowfly, Lucilia sericata, was the most common calliphorid trapped in these habitats. Neither of the two Lucilia species were found in native bush remnants in urban or rural areas where Calliphora hilli was dominant. Similarly very few L.cuprina (0.3% of the total) were trapped in rural rubbish tips in January and February where the majority of blowflies were again L.sericata. These results are compared with data collected from pastoral habitats, where L.cuprina is the major cause of flystrike. Lucilia cuprina was reared from five out of ninety-nine carcases found in rural areas. Calliphora stygia and L.sericata were the most common blowflies colonizing carrion and were reared from 59% and 51% of the carcases respectively.     

Bimodal cue complex signifies suitable oviposition sites to gravid females of the common green bottle fly

Gravid females of the common green bottle fly, Lucilia sericata Meigen (Diptera: Calliphoridae), readily locate recently deceased vertebrates as oviposition sites, particularly when these animals have been injured. We investigated semiochemical and visual cues that mediate attraction of gravid females to fresh rat carrion. Female flies were more strongly attracted to incised rat carrion than to intact carrion. They were also attracted to Porapak Q headspace volatile (HSV) extract of incised rat carrion. Analyzing aliquots of Porapak Q HSV extract by gas chromatographic‐electroantennographic detection revealed nine components [phenol, para‐ and/or meta‐cresol (could not be separated), guaiacol, dimethyl trisulfide (DMTS), phenylacetaldehyde, (E)‐2‐octenal, nonanal, and tetramethyl pyrazine] that consistently elicited responses from blow fly antennae. In laboratory experiments, a synthetic blend of these nine components was as attractive to gravid females as Porapak Q HSV extract, but blend attractiveness was due entirely to DMTS. In both laboratory and field experiments, increasing doses of DMTS attracted increasingly more flies. Coupled with DMTS, carrion‐type color cues (dark red, black) were more effective than bright color cues (white, yellow) in attracting flies. In field experiments, dark traps baited with DMTS captured a total of 214 calliphorid flies (200 L. sericata, 10 Lucilia illustris Meigen, three Calliphora vicina Robineau‐Desvoidy, one Calliphora vomitoria L.), all of which were gravid females. These results support the conclusion that DMTS and dark color represent a bimodal cue complex that signifies suitable oviposition sites to gravid calliphorid females, particularly L. sericata.     

Insect growth regulator cross-resistance studies in field- and laboratory-selected strains of the Australian sheep blowfly, Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae)

Abstract Resistance to diflubenzuron in the Australian sheep blowfly, Lucilia cuprina , has rendered this insecticide incapable of preventing flystrike in sheep from a few districts in eastern Australia. Wool producers affected by this situation must find suitable alternatives to protect their flocks. Results of laboratory bioassays against one population demonstrated that, despite extremely high diflubenzuron resistance (Resistance Factor >791), it had only very low (2x) tolerance of cyromazine and dicyclanil. It is unlikely that this level of tolerance would have any practical impact on field control with either insecticide. Consequently, wool producers in districts where diflubenzuron-resistant flies are common can rotate insecticide treatment to either of these compounds to prevent flystrike in their flocks. However, unlike the highly diflubenzuron-resistant field strain, a laboratory strain selected for resistance to diflubenzuron (Resistance Factor = 617) was 10 times more resistant to dicyclanil than a susceptible strain but, like the field strain, was only two times more tolerant of cyromazine. Conversely, a field-derived strain selected in the laboratory for cyromazine resistance was 20 times more resistant to dicyclanil and 362 times more resistant to diflubenzuron than the reference susceptible strain.     

Addressing pain caused by mulesing in sheep

The surgical operation of mulesing cuts wool-bearing and wrinkled skin from the perineal region and adjoining hindquarters of Australian Merino sheep, and has been shown in combination with tail docking to provide significantly enhanced protection against flystrike for the remainder of the animal's life. There is strong evidence that mulesing is a painful procedure, and its medium term future is doubtful in the face of animal welfare campaigns and societal expectations. Post-surgery analgesia for mulesing is available in the form of a topical anaesthetic treatment that provides amelioration of animal behavioural and physiological responses for some hours after the procedure. However, the topical anaesthetic alone does not provide analgesia during the operation, and its duration of action is less than the indicative period of pain and discomfort that follows mulesing. Greater analgesic coverage of mulesing-induced pain would appear to require the use of a non-steroidal anti-inflammatory drug such as carprofen in combination with topical anaesthesia, but most attention on addressing the mulesing issue is now focused on the development of non-surgical mulesing alternatives, breeding sheep that do not require mulesing, and targeting the fly and its larvae through an integrated pest management (IPM) approach. Non-surgical approaches to achieving a similar conformation change to that induced by mulesing have included research on topical and intradermal administration of chemicals and bioactive agents to cause skin scarring, stretching and depilation. Agents incorporating collagenase and quaternary ammonium compounds have been evaluated and rejected, including on animal welfare grounds. Intradermal sodium lauryl sulfate administration appears to induce a relatively mild behavioural and physiological response, but further evaluation and efficacy testing are required. A physical treatment aimed at achieving a similar result to surgical mulesing involves the placement of clips to remove clamped skin folds through ischaemia. The clip treatment is commercially available, and has been shown to induce only minimal animal discomfort and inflammatory responses. Early efficacy research suggests that breech flystrike rates may not be substantially improved by the clip procedure, and its viability as a long-term replacement for mulesing remains to be determined. The availability of chemical preventative treatments that provide sheep with up to 4 months’ protection against flystrike, combined with strategies such as increased crutching (shearing dirty wool off the breech), would allow many producers to cease mulesing. However, this would be at a greater cost and with the likely exit of a proportion from the industry, unless wool value reflected a substantive price differential for non-mulesed flocks. The genetic selection of Merino sheep that do not require post-birth modification to their breech skin, in combination with a strategic IPM program, would appear to provide the most sustainable approach to managing breech flystrike in the Australian Merino. Such a breeding program requires a sustained commitment over at least a 10-year period, and is not helped by attitudes of some within the industry that conventional mulesing does not have a finite lifespan as a flystrike control option.     

Volatiles released from foliar extract of host plant enhance landing rates of gravid Polygonia c‐album females,but do not stimulate oviposition

The role of olfactory cues for host search is much less investigated in day‐active butterflies than in their relatives, the nocturnal moths. The goal of this study was to investigate whether host‐plant volatiles from foliar extracts of hop, Humulus lupulus L. (Cannabaceae), evoke electroantennographic (EAG) responses, increase landing rates, and stimulate egg‐laying behavior of gravid Polygonia c‐album L. (Lepidoptera: Nymphalidae) females. Eighty‐nine volatile compounds were detected in a non‐concentrated methanol extract of hop by gas chromatography–mass spectrometry, 11 of which elicited an EAG response. Concentration of the crude extract significantly reduced landing rates on artificial leaves treated with the sample due to loss of volatile compounds, but after landing the oviposition response of gravid females was not affected. A mixture of eight commercially available EAG‐active volatiles increased the landing rate of gravid females to their source but did not act as oviposition stimulants. Dividing the volatile compounds into two groups – consisting of (1) hexanal, (E)‐2‐hexenal, octanal, nonanal, and decanal, and (2) sulcatone, humulene, and benzyl alcohol – obliterated effectiveness, revealing synergism between compounds. Although volatiles did not stimulate oviposition, they significantly contributed to the distribution of eggs by increasing the landing rates on treated artificial leaves.     

Attraction of the sandfly Lutzomyia longipalpis to possible biomarker compounds from dogs infected with Leishmania infantum

Lutzomyia longipalpis (Diptera: Psychodidae) is the primary vector of Leishmania infantum (Kinetoplastida: Trypanosomatidae) in the Americas. Studies have been carried out to identify new alternatives for monitoring and controlling this sandfly species, particularly with the use of chemical baits. The attractiveness of odours emitted by foxes and alcohols found in some plants has already been demonstrated in laboratory tests with Lu. longipalpis. However, no studies have evaluated the responses of these insects to volatile organic compounds (VOCs) emitted by dogs. The present study was carried out to investigate the effects on Lu. longipalpis of individual and blends of VOCs identified in hair from dogs infected with L. infantum. Effects in male and female Lu. longipalpis were assessed using wind tunnel methodology. Individual compounds including octanal, nonanal, decanal and heptadecane showed capacity for activating and/or attracting male Lu. longipalpis. Only decanal and nonanal showed effects on females. The combination of octanal, decanal and heptadecane increased activation and attraction behaviour in male sandflies, as did the blend of octanal and decanal. These findings indicate that VOCs emitted by dogs may be an interesting source of new attractants of sandflies.     

Cloning,recombinant expression and inhibitor profiles of dihydrofolate reductase from the Australian sheep blow fly,Lucilia cuprina

While dihydrofolate reductase (DHFR) is an important drug target in mammals, bacteria and protozoa, no inhibitors of this enzyme have been developed as commercial insecticides. We therefore examined the potential of this enzyme as a drug target in an important ectoparasite of livestock, the Australian sheep blow fly, Lucilia cuprina (Diptera: Calliphoridae) (Wiedemann). The non‐specific DHFR inhibitors aminopterin and methotrexate significantly inhibited the growth of L. cuprina larvae, with IC₅₀ values at µg levels. Trimethoprim and pyrimethamine were 5–30‐fold less active. Relative IC₅₀ values for the inhibition of recombinant L. cuprina DHFR by various inhibitors were in accordance with their relative effects on larval growth. The active‐site amino acid residues of L. cuprina DHFR differed by between 34% and 50% when compared with two mammalian species, as well as two bacteria and two protozoa. There were significant charge and size differences in specific residues between the blow fly and human DHFR enzymes, notably the L. cuprina Asn21, Lys31 and Lys63 residues. This study provides bioassay evidence to highlight the potential of blow fly DHFR as an insecticide target, and describes differences in active site residues between blow flies and other organisms which could be exploited in the design of blow fly control chemicals.     

Insecticide resistance and malathion carboxylesterase in the sheep blowfly,Lucilia cuprina

Resistance to the organophosphorus insecticide malathion in genetically related strains of the Australian sheep blowflyLucilia curprina was examined. Separate lines of blowflies were established by homozygosis of the fourth chromosome of the parental RM strain. Both the RM and the derived resistant (der-R) strains are approximately 100 times more resistant to malathion than the related susceptible der-S strain, resistance being correlated with a 45- to 50-fold increase in a malathion carboxylesterase (MCE) activity. MCE has a pH optimum ranging between 6.6 and 8.0 and is strongly inhibited by the carboxylesterase inhibitors triphenyl phosphate, paraoxon, and diiospropylfluorophosphate. Subcellular fractionation revealed that MCE was localized predominantly to the cytosol and mitochondria in both resistant and susceptible blowflies. A single MCE was purified to homogeneity from RM blowflies. It has a pI of 5.5, is a monomer of 60.5 kDa, and hydrolyzes malathion with aV _max of 755 nmol/min/mg protein and aK _m of 11.0 µM. L. cuprina have thus evolved a remarkable MCE which is faster and more efficient at hydrolyzing a specific insecticide than any other insect esterase yet described.     

Influence of olfactory stimulants on resistance/susceptibility of pearl millet, Pennisetum americanum to the rice weevil, Sitophilus oryzae

The orientation responses of the rice weevil, Sitophilus oryzae (L.) (Coleoptera: Curculionidae) as influenced by the olfactory stimulants emanating from either resistant or susceptible pearl millet, Pennisetum americanum, kernels were studied using a Y-tube olfactometer. The rice weevils were more attracted towards odours from millet grains than to blank controls and to susceptible entries more than to resistant entries, in general. However, there were exceptions. Odours from some resistant entries caused statistically the same, or even more positive orientations than those from some susceptible entries, and this suggests that the relative resistance of an entry and its olfactory attractiveness may not be strongly related, or that the resistance to infestation might be influenced by factors other than attractiveness. The partial damage of the millet kernel pericarp by abrasion slightly increased the orientational response of the weevils to odours from some susceptible and some resistant entries alike.     

Evaluation of reference genes for real‐time PCR quantification of gene expression in the Australian sheep blowfly,Lucilia cuprina

The Australian sheep blowfly, Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae), is an important pest of sheep in Australia and other parts of the world. However, the paucity of information on many fundamental molecular aspects of this species limits the ability to exploit functional genomics techniques for the discovery of new drug targets for its control. The present study aimed to facilitate gene expression studies in this species by identifying the most suitable reference genes for normalization of mRNA expression data. Quantitative real‐time polymerase chain reaction (PCR) was performed with 11 genes across RNA samples from eggs, L1, L3, pupae and adult life stages, and two normalization programs, Normfinder and geNorm, were then applied to the data. The results showed an ideal set of genes (18S rRNA, 28S rRNA, GST1, β‐tubulin and RPLPO) for data normalization across all life stages. The most suitable reference genes for studies within specific life stages were also identified. GAPDH was shown to be a poor reference gene. The reference gene recommendations in this study will be of use to laboratories investigating gene expression in L. cuprina and related blowfly species     

Electroantennographic responses of red flour beetle Tribolium castaneum Herbst (Coleoptera: Tenebrionidae) to volatile organic compounds

The red flour beetle Tribolium castaneum Herbst (Coleoptera: Tenebrionidae) is the most encountered and destructive stored product insect pest of cereal grains and seeds. Although this beetle has been used as a model organism for many decades, there is no systematic knowledge about antennal detection of host and non‐host volatiles. Electroantennogram responses to 94 selected volatile organic compounds including alkanes, alkenes, alcohols, aldehydes, ketones, carboxylic acids, esters, terpenoids and aromatic compounds were recorded from both sexes of T. castaneum. Overall, female and male T. castaneum exhibited similar electroantennography (EAG) responses. Compounds eliciting the strongest EAG responses within compound groups of chemical similarity were undecane, 1‐hexen‐3‐ol, octanal, 2‐heptanone, hexanoic acid and ethyl hexanoate. Comparison of vapour pressure and EAG amplitudes within homologous series of compounds revealed responses to undecane, octadecane, octanal, nonanal, 2‐heptanone, hexanoic acid and octanoic acid as outstanding. Given that systematic EAG screenings have not been conducted before, these are the best candidates for evaluation in future behavioural studies to unravel their potential for application in integrated pest management strategies of T. castaneum.     

Antennal and behavioural responses of Heortia vitessoides females to host plant volatiles of Aquilaria sinensis

This study investigated volatiles from Aquilaria sinensis (Lour.) Gilg (Thymelaeaceae) leaves that attracted Heortia vitessoides Moore (Lepidoptera: Crambidae). Volatiles from young and old A. sinensis leaves were identified and quantified by gas chromatography–mass spectrometry and gas chromatography‐electroantennogram detection. Both wind tunnel bioassays and field tests were conducted to measure the attraction of adults to synthetic blends of volatiles from leaves of different maturations. Consistent electroantennographic activity was obtained for nine and three compounds from headspace collections of young and old A. sinensis leaves, respectively. Qualitative and quantitative differences were found among two odour profiles. In wind tunnel experiments, the fresh young leaves proved to be more attractive to females than old leaves. A nine‐component mixture, including hexanal, limonene, 2‐hexanol, octanal, (Z)‐3‐hexenyl acetate, (Z)‐3‐hexen‐1‐ol, nonanal, decanal, and 2,6,10‐trimethyl‐dodecane (with a ratio of 2:16:9:4:63:100:13:10:5) from young leaves attracted moths significantly more than the three‐component mixture of nonanal, decanal, and 2,6,10‐trimethyl‐dodecane (with a ratio of 11:14:26) from old leaves. Further subtractive bioassays conducted in the wind tunnel showed that both the complete nine‐component mixture and a subtracted four‐component mixture of hexanal, (Z)‐3‐hexenyl acetate, nonanal, and decanal (with a ratio of 2:63:13:10) elicited equivalent responses in females. All components in the four‐component blend were essential for optimal attraction. In a field trial using the nine‐ and four‐component blends, more moths were captured using both blends than in traps baited with hexane only. Our study indicates that the odour blends of young leaves play an important role in H. vitessoides host plant recognition. The mechanisms behind host recognition and age‐dependent changes in leaf chemistry are discussed.     

Responses of <Emphasis Type="Italic">Ceutorhynchus obstrictus</Emphasis> (Marsham) (Coleoptera: Curculionidae) to olfactory cues associated with novel genotypes developed by <Emphasis Type="Italic">Sinapis alba</Emphasis> L. × <Emphasis Type="Italic">Brassica napus</Emphasis> L.

Many herbivorous insects use olfactory cues for host location. Extracts from Brassica napus L. have been shown to elicit electrophysiological and behavioural responses in the cabbage seedpod weevil, Ceutorhynchus obstrictus (Marsham) (syn. C. assimilis (Paykull)) (Coleoptera: Curculionidae). These include volatile products of the hydrolysis of glucosinolates. Here we present results of a laboratory olfactometer study examining the attractiveness of odours from flowering racemes and foliage of Sinapis alba L. (an inappropriate host for larval development), B. napus (an excellent host for larval development) and lines derived from S. alba × B. napus selected from colonization studies to demonstrate resistance or susceptibility. Results of this study indicate differential attraction of C. obstrictus to the odours of resistant and susceptible lines and suggest the role of hydrolysis products of glucosinolates, particularly the attractive effects of 2-phenylethyl isothiocyanate.     

Organisation and expression of a cluster of yolk protein genes in the Australian sheep blowfly, Lucilia cuprina

The Australian sheep blowfly Lucilia cuprina is a major pest for the Australian and New Zealand sheep industries. With the long-term aim of making a strain of L. cuprina suitable for a genetic control program, we previously developed a tetracycline-repressible female lethal genetic system in Drosophila. A key part of this system is a female-specific promoter from a yolk protein (yp) gene controlling expression of the tetracycline-dependent transactivator (tTA). Here we report the sequence of a 14.2 kb genomic clone from L. cuprina that contains a cluster of three complete yp genes and one partial yp gene. The Lcyp genes are specifically expressed in females that have received a protein meal. A bioinformatic analysis of the promoter of one of the yp genes (LcypA) identified several putative binding sites for DSX, a known regulator of yp gene expression in other Diptera. A transgenic strain of L. cuprina was made that contained the LcypA promoter driving the expression of the Escherichia coli lacZ reporter gene. Transgenic females express high levels of β-galactosidase after a protein meal. Thus the LcypA promoter could be used to obtain female-specific expression of tTA in transgenic L. cuprina.     

Electrophysiological and behavioural responses of mosquitoes to volatiles of Silene otites (Caryophyllaceae)

In order to understand the biological significance of flower odour for attraction of mosquitoes, electrophysiological responses to headspace flower odour samples of Silene otites (L.) Wibel were investigated on Culex pipiens pipiens biotype molestus Forskal 1775 and Aedes aegypti L. using coupled gas chromatographic-electroantennographic detection (GC-EAD). No remarkable differences in antennal responses to the odour compounds have been found between these two mosquito species. Further, the behavioural attractiveness of the electrophysiologically active compounds, singly or as multiple odour mixtures, was evaluated with bioassay experiments with C. pipiens molestus. In bioassays, C. pipiens responded to 14 electrophysiologically active compounds in different magnitudes (65–20%) and acetophenone, linalool oxide (pyranoid), phenyl acetaldehyde and phenylethyl alcohol were found as more attractive in comparison to the least attractive compound, hexanol. In two-stimulus choice test, mosquitoes were significantly more attracted to the mixture of the four most attractive compounds compared to the mixture of all 14 compounds. The results of present study confirm that floral odours are attractive cues for mosquitoes.     

In vitro antibacterial activity and physicochemical properties of a crude methanol extract of the larvae of the blow fly Lucilia cuprina

The emergence of multidrug‐resistant bacterial strains has prompted the reintroduction of maggot therapy in the treatment of chronic, infected wounds. Many previous studies have demonstrated the potent antibacterial activity of larval excretions/secretions of the blowfly Lucilia sericata (Meigen) (Diptera:Calliphoridae) against bacteria. However, the antibacterial activity of its sibling species, Lucilia cuprina (Wiedemann) (Diptera:Calliphoridae) against a wide range of pathogenic bacteria has never been determined. The aim of this study was to develop a new procedure to produce whole body extract of larvae of L. cuprina via methanol extraction as well as to demonstrate the in vitro antibacterial activity of this extract against seven selected wound pathogens (Staphylococcus aureus, methicillin‐resistant S. aureus, S. epidermidis, Streptococcus pyogenes, Klebsiella pneumoniae, Pseudomonas aeruginosa and Escherichia coli). The turbidimetric assay demonstrated that L. cuprina larval extract was significantly potent against all bacteria tested (P < 0.001). Additionally, colony‐forming unit (CFU), agar well diffusion and minimum inhibitory concentration assays have confirmed the apparent potency of larval extract against P. aeruginosa. The reconstituted larval extract was highly robust and thermally stable. These observations substantiated the feasibility of the methanol extraction method in the production of larval extract.     

The effects on genetic variability following a recent colonization event: the Australian sheep blowfly, Lucilia cuprina arrives in New Zealand

The Australian sheep blowfly, Lucilia cuprina, was first identified in New Zealand in 1988 and is now found to have spread throughout many sheep-farming regions. L. cuprina is estimated to have been present in New Zealand < 20 years, while in Australia L. cuprina has been estimated present > 100 years. The aim of this study was to determine the genetic effects of colonization of L. cuprina and to compare populations of L. cuprina from these two countries in terms of genetic variability and differentiation. Allozyme electrophoresis was used which revealed variability at six loci. 1680 blowflies were examined from 56 sites throughout L. cuprina's range in both countries. Genetic variability at each locus in terms of allele composition was found to be high and genetic differentiation varied considerably in New Zealand in comparison to Australia. Temporal sampling in New Zealand suggests seasonal fluctuations of population size in the recently colonized region of the South Island.     

Spectrum of cross-resistance to insecticides in field samples of the primary sheep blowfly, Lucilia cuprina

Hughes P. B. (1981) Spectrum of cross-resistance to insecticides in field samples of the primary sheep blowfly, Lucilia cuprina. Internationaljournal for Parasitology11:475–479. Resistance to the organophosphorous (OP) insecticide, diazinon, is due to a major gene on chromosome 4. To examine the spectrum of cross-resistance in field populations of Lucilia cuprina, larvae from 41 field samples were exposed to the susceptible discriminating doses of five insecticides. A comparison of the mortalities produced by diazinon to those caused by the other OPs tested (fenthion-ethyl, chlorfenvinphos and coumaphos) indicates that the diazinon resistance gene may confer cross-resistance to other OPs used for blowfly control. However, two samples were found to be resistant to diazinon but susceptible to coumaphos. The OP resistance gene is present at a high frequency in field populations, with a mean frequency of 0–82. Mortalities produced by the carbamate, butacarb, did not relate to those due to diazinon. It seems that a separate mechanism for carbamate resistance is also present in the field. No indication of resistance was found to the insect growth regulator Vetrazin^® in any sample tested.     

Is aggregated oviposition by the blow flies Lucilia sericata and Phormia regina (Diptera: Calliphoridae) really pheromone‐mediated?

When female blow flies Lucilia sericata and Phormia regina (Diptera: Calliphoridae) oviposit in aggregations on carrion, even‐aged larval offspring reportedly develop faster, and fewer are parasitized or preyed upon. The benefits of aggregated oviposition equally affect con‐ and heterospecific larvae sharing a resource. The benefits imply that female blow flies engage in coordinated, pheromone‐mediated oviposition behavior. Yet, repeated attempts to identify oviposition pheromones have failed invoking doubt that they exist. Simply by regurgitating and feeding on carrion, flies may produce attractive semiochemicals. If flies were to aggregate in response to feeding flies rather than ovipositing flies, then the semiochemical cue(s) may be associated with the salivary gland. Working with L. sericata and P. regina and using liver as a surrogate oviposition medium, we test the hypotheses, and present data in their support, that (i) gravid or nongravid females ovipositing and/or feeding on liver enhance its attractiveness to gravid and nongravid females; (ii) females respond to semiochemicals from feeding heterospecific females; (iii) females respond equally well to semiochemicals from feeding con‐ and heterospecific females; (iv) macerated head tissues of females applied to liver enhance its attractiveness; and (v) females in direct contact with and feeding on liver, but not when next to yet physically separated from liver, enhance attraction of flies. We conclude that oviposition site‐seeking females do not respond to an oviposition pheromone. Instead, they appear to coopt semiochemicals associated with feeding flies as resource indicators, taking chances that resources are suitable for oviposition, and that ovipositing flies are present.