A comparison of Lucitraps and sticky targets for sampling the blowfly Lucilia sericata

The Lucitrap (Miazma Pty Ltd, Queensland, Australia) combined with a synthetic odour bait, Lucilure (Miazma Pty Ltd, Queensland, Australia), is a commercially available trap for sampling and control of Lucilia cuprina (Wiedemann) in Australia. It was tested in Hungary against Lucilia sericata (Meigen) (Diptera: Calliphoridae), a cause of sheep strike throughout temperate Europe. The standard Lucitrap was tested against black or yellow sticky target traps. Both trap types were baited with either Lucilure or liver and 10% w/v sodium sulphide solution. With Lucilure as bait, L. sericata were caught on sticky traps but not in Lucitraps. With liver and sodium sulphide as bait, sticky traps caught 500-1500 times more L. sericata than Lucitraps. An adhesive sheet fitted to the top of a Lucitrap captured 30-300 times more L. sericata then were captured inside an unaltered Lucitrap. Direct observation of metallic green calliphorids (92.1% L. sericata) alighting on Lucitraps indicated that most flies stayed for a short while (modal class 2-4 s) and only a few stayed longer, to an observed maximum of 28 s. Flies explored a mean of 1.5 entry holes (range 0-7) during a visit but only 6% entered the trap. Size of L. sericata was not a physical barrier to Lucitrap entry, because many larger species were captured. However, L. sericata captured inside Lucitraps were significantly smaller than those captured on sticky traps, demonstrating that size was of behavioural importance. The data demonstrate that the Lucitrap is not effective as a trap for L. sericata in Hungary, due mainly to a failure of flies to enter the trap in large numbers. In Australia and South Africa, L. sericata is commonly caught in Lucitraps baited with Lucilure, although L. cuprina is more numerous. Our study highlights the potential for diversity of fly behaviour between different geographical populations of the same species. Such diversity can have a significant effect on the functioning of systems for fly sampling and control, when these systems depend for their success on certain behavioural responses of the target species.     

mtDNA中COⅠ分子标记在常见食尸性蝇类鉴定中的应用

死后不同时间,在尸体上出现不同种类食尸性蝇类的演替规律,可用于准确推断死亡时间。传统上仅依据蝇类形态学特征来判断种属,但由于蝇类的形态结构复杂和种间形态差异微小等特点,对蝇类尤其是对蝇类幼虫的种属鉴别很难。因此应用分子生物学方法对食尸性蝇类及其幼虫进行种属鉴定非常重要。本研究主要是利用此方法对我国西部部分地区常见双翅目食尸性蝇类包括：开普黑蝇、大头金蝇、丝光绿蝇及部分卵,铜绿蝇、棕尾别麻蝇及部分幼虫和蛹的线粒体DNA(mtDNA)上细胞色素氧化酶辅酶Ⅰ(COⅠ)中278 bp的基因序列进行鉴别。除个别蝇类如丝光绿蝇与铜绿蝇外,该方法均能有效地将上述食尸性蝇类鉴定到种属水平。在我国,它将成为法医鉴别食尸性蝇类种属的可靠依据。     

Analysis of the circadian clock gene period in the sheep blow fly Lucilia cuprina

We have isolated a homologue of the period (per) gene from the Australian sheep blow fly, Lucilia cuprina, as part of a comparative approach to the analysis of dipteran circadian systems. Sequence analysis of the 4 kb per cDNA revealed the conservation of three functional domains, namely the PAS dimerization motif, and the nuclear and cytoplasmic localization domains. A fourth domain, the threonine-glycine (TG) repeat region, is also conserved in L. cuprina per but has been severely truncated. No length variation was found in the TG repeat of L. cuprina or L. sericata collected from several different latitudinal zones. Expression analysis indicated a diel oscillation in per mRNA in LD 12:12 with a period of 24 h and a peak at Zt 12. PER-immunoreactive protein oscillations were also demonstrated, with peak immunoreactivity lagging approximately 3 h behind peak mRNA levels. These results show the existence of a Drosophila-like circadian system in a calliphorid fly. They also provide evidence for the conservation of per function across the Diptera, and confirm the relevance of the Drosophila system as a model for fly circadian rhythms.     

Suppression of populations of Australian sheep blowfly, Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae), with a novel blowfly trap

The Australian sheep blowfly, Lucilia cuprina , initiates more than 85% of fly strikes on sheep in Australia with an estimated average annual cost of A$280 million to the Australian sheep industry. LuciTrap^® is a commercially available, selective trap for L. cuprina consisting of a plastic bucket with multiple fly entry cones and a synthetic attractant. The impact of LuciTrap on populations of L. cuprina on sheep properties in five Australian states was evaluated by comparing L. cuprina populations on paired properties with and without LuciTraps over seasons when significant fly populations could be expected. Twenty-four comparisons (trials) were conducted over 4 years. During times of 'higher fly density' (when the 48 h geometric mean of trap catches on the control property was greater than five L. cuprina ), the overall geometric mean trap catches for control and trapped properties differed significantly ( P  < 0.001) with mean trap catches of 19.4 and 7.74 L. cuprina , respectively. The selectivity of the LuciTrap was confirmed with 59% of all trapped flies being L. cuprina . Chrysomya spp. and Calliphora spp. constituted 9.3% and 1.1% of the catches with a variety of other flies (mainly Sarcophagidae and Muscidae ) providing the remainder (31%). Lucilia sericata was only trapped in Tasmania and made up 7.7% of the Lucilia spp. catch in that state. Seventy-two per cent of the trapped L. cuprina were female. The deployment of LuciTrap on sheep properties at one trap per 100 sheep from the beginning of the anticipated fly season suppressed the populations of L. cuprina by 60% compared with matched control properties. The LuciTrap is a selective and easy to use fly trap and constitutes an effective, non-insecticidal tool for use in integrated management programs for L. cuprina .     

LABORATORY STUDIES ON THE CHEMO-SENSORY REACTIONS OF BLOWFLIES

Attraction of various blowflies to sheep wool has been demonstrated under laboratory conditions using a choice-chamber technique. The attractive factor in wool is intimately associated with the wool fibre for it does not disappear during storage and it is not completely removed by washing.
Only female flies were attracted to wool. The highest degree of attraction was obtained from fertilized females which had daily access to meat. Even in the absence of fertilization and meat, attraction was not completely abolished.
The species and strains tested can be placed in the following order in terms of strength of response to sheep wool: Lucilia sericata (British), L. cuprina (tested on damp floor)>L. sericata (Danish: country)> L. sericata (Danish: city), L. sericata (Australian) > L. illustris, L. caesar. No response was given by Calliphora vomitoria.
The olfactory organs involved in the response to wool are mainly on the antennae but antennaeless L. sericata and L. cuprina with some tarsi removed were still able to show some response to wool provided that they were tested on a damp floor. In the course of testing the olfactory responses it was also shown that the main humidity receptors are present on the antennae.
The existence of a 'wool-factor' to which only certain strains of L. sericata and L. cuprina are specially sensitive offers a possible explanation for the greater efficiency of these species as myiasis-producing forms.     

Blowfly succession from possum (Trichosurus vulpecula) carrion in a sheep-farming zone

The significance of brushtail possum, Trichosurus vulpecula Kerr (Diprotodontia: Phalangeridae) carcasses to the succession and production of Diptera species and its relevance to fly strike management in Tasmania, Australia was examined. Calliphora stygia (Fabricius), Lucilia sericata (Meigen) and Calliphora vicina Robineau-Desvoidy (Diptera: Calliphoridae) were found to be the most abundant and Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae) always the least abundant (< 1%) of the putative primary fly invading species to emerge. Carcasses that were left for up to 15 days in the field before being exposed to flies for 2 days also acted as breeding sites for large numbers of all primary fly species, with the exception of L. cuprina. Ordination analysis revealed no relationship between possum carcasses according to their length of exposure but did show significant negative associations between the number of putative secondary invaders (Chrysomya rufifacies (Macquart) (Diptera: Calliphoridae), Chrysomya varipes (Macquart) (Diptera: Calliphoridae) and putative tertiary flies (Hydrotaea rostrata Robineau-Desvoidy (Diptera: Muscidae)) to the number of C. vicina or C. stygia to emerge. There was enormous variability in the numbers of secondary/tertiary fly species to emerge from carcasses (0-11 450) that negatively correlated with the proportion of all flies to emerge that were primary, and with the mean size of adult L. sericata. Although carcass temperatures, especially those with a large larval population, were elevated, this did not appear to result in significant pre-adult fly mortality. The most important primary fly strike species L. cuprina was only found in insignificant numbers, whereas three other members of the fly strike fauna C. stygia, L. sericata and Ch. rufifacies did use possum carrion as an important breeding resource, but left implications for fly strike management inconclusive.     

The use of carrion as breeding sites by the blowfly Lucilia sericata and other Calliphoridae

Abstract. To examine the species composition of flies breeding in carrion in the field, the carcasses of mice and quail were exposed on sheep farms in the South West of England. Calliphora vicina was the dominant species of Diptera; 19,294 individuals emerged from 175 of the 241 infested carcasses recovered. Lucilia sericata emerged from only 39 of the infested carcasses, at a median of 10 adults per infested carcass. Other species of Lucilia present were L.ampullacea, L.caesar and L.illustris. The highest number of L.sericata emerged from carcasses placed in open pasture, the highest number of C.vicina emerged from carcasses in hedgerow, whereas the highest numbers of L.caesar, L.ampullacea and L.illustris emerged from carcasses in woodland. The duration of exposure of carcasses in the field was negatively related to the size of the adult L.sericata which emerged and, in woodland and hedgerow habitats, to the number of L.sericata which emerged. These data indicate that the larvae of L.sericata in carcasses experience significant levels of competition and that the intensity of this competition may be sufficient to reduce the numbers of L.sericata able to emerge successfully. The size distributions of female L. sericata which emerged from carcasses or which were caught as adults in the field showed only a small degree of overlap, suggesting that only a relatively small proportion of the wild L.sericata population emerge from carcass breeding sites. The results are discussed in relation to the development of new approaches to the control of blowfly strike of sheep.     

Cloacal myiasis by Lucilia spp. (Diptera: Calliphoridae) in a rooster (Gallus gallus domesticus) and two Harris's hawks (Parabuteo unicinctus)

In this study, cloacal myiasis caused by dipterans of Lucilia genus was found in a rooster (Gallus gallus domesticus) and two Harris's hawks (Parabuteo unicinctus) from Peru. Larval dipteran were collected and preserved in ethanol. Morphological analysis indicated two species: Lucilia sericata in the rooster and in one Harris's hawk, and Lucilia cuprina in the other Harris's hawk. Molecular analysis confirmed the diagnosis by amplification of the nucleotide sequences of the cytochrome c oxidase subunit 1 gene and internal transcribed spacer 2 region. The sequences were compared with sequence references from a public sequence database, which showed a 100% matched identity. This study demonstrated for first time cloacal myiasis by L. sericata in a domestic bird from Peru and in Harris's hawk. Also, for the first time, L. cuprina was found in a bird of prey.     

Mitochondrial genomes of the sheep blowfly, Lucilia sericata, and the secondary blowfly, Chrysomya megacephala

This paper presents complete mitochondrial genomes for the sheep blowfly, Lucilia sericata (Meigen), and the secondary blowfly, Chrysomya megacephala (Fabricius). Both L. sericata and C. megacephala had standard dipteran-type mitochondrial genome architectures and lengths of 15 945 bp and 15 831 bp, respectively. Additionally, C. megacephala possessed a tRNA duplication either side of the D-loop, as previously reported in another Chrysomya species, C. putoria; this duplication appears to be synapomorphic for the genus Chrysomya. As in other insect mitochondrial genomes, base compositions had a high AT content, with both genomes more than 76% AT-rich.     

The molecular systematics of blowflies and screwworm flies (Diptera: Calliphoridae) using 28S rRNA, COX1 and EF-1α: insights into the evolution of dipteran parasitism

The Calliphoridae include some of the most economically significant myiasis-causing flies in the world - blowflies and screwworm flies - with many being notorious for their parasitism of livestock. However, despite more than 50 years of research, key taxonomic relationships within the family remain unresolved. This study utilizes nucleotide sequence data from the protein-coding genes COX1 (mitochondrial) and EF1α (nuclear), and the 28S rRNA (nuclear) gene, from 57 blowfly taxa to improve resolution of key evolutionary relationships within the family Calliphoridae. Bayesian phylogenetic inference was carried out for each single-gene data set, demonstrating significant topological difference between the three gene trees. Nevertheless, all gene trees supported a Calliphorinae-Luciliinae subfamily sister-lineage, with respect to Chrysomyinae. In addition, this study also elucidates the taxonomic and evolutionary status of several less well-studied groups, including the genus Bengalia (either within Calliphoridae or as a separate sister-family), genus Onesia (as a sister-genera to, or sub-genera within, Calliphora), genus Dyscritomyia and Lucilia bufonivora, a specialised parasite of frogs and toads. The occurrence of cross-species hybridisation within Calliphoridae is also further explored, focusing on the two economically significant species Lucilia cuprina and Lucilia sericata. In summary, this study represents the most comprehensive molecular phylogenetic analysis of family Calliphoridae undertaken to date.     

Development of an attractive target for the sheep blowfly Lucilia sericata

Coloured, odour-baited, sticky targets, designed to catch the sheep blowfly Lucilia sericata Meigen are described and the results of field trials are presented. The number of L. sericata caught by the targets was positively related to target surface area and was dependent on target colour. The performance of the coloured targets is described by a model in which catch is related negatively to reflectivity in the 300-450 nm (ultra-violet/blue) band of the spectrum and positively to the reflectivity in the 450-580 nm (blue-green-yellow) band. Of the Lucilia blowflies caught by the targets, 13-16% were L.sericata and 72-75% of the L.sericata were female. The number of L.sericata caught appeared to be positively related to the availability of sheep over a relatively localized area and showed a marked decline following insecticidal dipping of sheep. The further development of targets and the implications of the results for L.sericata control are discussed.     

Responses of the sheep blowflies Lucilia sericata and Lcuprina to odour and the development of semiochemical baits

Abstract. The literature relating to the attraction of the sheep blowflies Lucilia sericata and Lucilia cuprina to their ovine hosts is reviewed. The responses of the two species are similar and different components of the behaviour leading to host location and oviposition appear to involve at least two distinct sets of semiochemical cues. Activation, upwind orientation and landing appear to occur in response to putrefactive sulphur-rich volatiles, originating from bacterial decomposition products. Oviposition is elicited primarily by the presence of ammonia-rich compounds; moisture, pheromones and tactile stimuli may also act as oviposition stimuli. There is a pronounced sex difference in the response of Lucilia to semiochemicals with a higher proportion of females attracted than males and a higher proportion of gravid than non-gravid females. While the mechanisms of host location by Lucilia are of intrinsic interest, understanding the responses to semiochemicals is important in the attempt to develop powerful synthetic baits for deployment with the traps or targets used for population sampling or suppression. The literature is discussed with respect to the development of synthetic semiochemical baits.     

Prasinons A and B: Potent Insecticides from Streptomyces prasinus

Two metabolites which have high activity against sheep blowfly larvae (Lucilia sericata and L. cuprina) were found to be produced by Streptomyces prasinus NCIB 10719. These substances were isolated from culture filtrate by solvent extraction and chromatography and named prasinons A and B. Fermentation factors affecting the formation of these substances are described together with their physical, chemical, and biological properties.     

The acetylcholinesterase gene and organophosphorus resistance in the Australian sheep blowfly, Lucilia cuprina

Acetylcholinesterase (AChE), encoded by the Ace gene, is the primary target of organophosphorous (OP) and carbamate insecticides. Ace mutations have been identified in OP resistants strains of Drosophila melanogaster. However, in the Australian sheep blowfly, Lucilia cuprina, resistance in field and laboratory generated strains is determined by point mutations in the Rop-1 gene, which encodes a carboxylesterase, E3. To investigate the apparent bias for the Rop-1/E3 mechanism in the evolution of OP resistance in L. cuprina, we have cloned the Ace gene from this species and characterized its product. Southern hybridization indicates the existence of a single Ace gene in L. cuprina. The amino acid sequence of L. cuprina AChE shares 85.3% identity with D. melanogaster and 92.4% with Musca domestica AChE. Five point mutations in Ace associated with reduced sensitivity to OP insecticides have been previously detected in resistant strains of D. melanogaster. These residues are identical in susceptible strains of D. melanogaster and L. cuprina, although different codons are used. Each of the amino acid substitutions that confer OP resistance in D. melanogaster could also occur in L. cuprina by a single non-synonymous substitution. These data suggest that the resistance mechanism used in L. cuprina is determined by factors other than codon bias. The same point mutations, singly and in combination, were introduced into the Ace gene of L. cuprina by site-directed mutagenesis and the resulting AChE enzymes expressed using a baculovirus system to characterise their kinetic properties and interactions with OP insecticides. The K(m) of wild type AChE for acetylthiocholine (ASCh) is 23.13 microM and the point mutations change the affinity to the substrate. The turnover number of Lucilia AChE for ASCh was estimated to be 1.27x10(3) min(-1), similar to Drosophila or housefly AChE. The single amino acid replacements reduce the affinities of the AChE for OPs and give up to 8.7-fold OP insensitivity, while combined mutations give up to 35-fold insensitivity. However, other published studies indicate these same mutations yield higher levels of OP insensitivity in D. melanogaster and A. aegypti. The inhibition data indicate that the wild type form of AChE of L. cuprina is 12.4-fold less sensitive to OP inhibition than the susceptible form of E3, suggesting that the carboxylesterases may have a role in the protection of AChE via a sequestration mechanism. This provides a possible explanation for the bias towards the evolution of resistance via the Rop-1/E3 mechanism in L. cuprina.     

Gold labelling of RNA in virus-induced mitochondrial vesicles in the sheep blowfly Lucilia cuprina.

Membranous vesicles are thought to be the replication site for viral RNA of many plant and animal viruses. A relatively rare site of virus-associated vesiculation is that of the mitochondrial outer membrane. In this study, virus-induced mitochondrial vesicles of the blowfly, Lucilia cuprina, were labelled with ribonuclease/gold and an antibody against double stranded RNA (anti-polyinosinic:polycytidylic acid). Both methods showed the presence of RNA in the vesicles thus indicating they may be a site for viral RNA replication in Lucilia.     

Salivary gland chromosomes of Lucilia cuprina (Diptera, Caliphoridae).

The salivary gland chromosome complement in Lucilia cuprina consists of five pairs of autosomes and two sex chromosomes X and Y, all with characteristic banding patterns. A standard map for the salivary gland chromosomes of this species has been prepared, and compared with the known map of Lucilia cuprina dorsalis from Australia. The differences in their banding patterns indicate the existence of different varieties within the same species.     

The amino acid sequence of cytochrome c from the blowfly Lucilia cuprina.

The amino acid sequence of cytochrome c isolated from the sheep blowfly Lucilia cuprina has been determined by comparison of the compositions of the tryptic peptides to those predicted from the published sequences of cytochromes c from other insects. Cytochrome c from L. cuprina differs at a single residue when compared to cytochrome c from the screw worm fly Haematobia irritans, a species belonging to the same order as the blowfly. This substitution, proline for alanine, has been located at position 44 in the protein chain.     

Progress towards the development of a transgenic strain of the Australian sheep blowfly (Lucilia cuprina) suitable for a male-only sterile release program

The Australian sheep blowfly Lucilia cuprina is the most important pest species involved in cutaneous myiasis (flystrike) of sheep in Australia and New Zealand. In New Zealand L. cuprina is primarily controlled through the application of insecticides. However, there is an increased interest in biological methods of control of this species. We have proposed to develop a genetically modified strain of L. cuprina that would be ideal for a male-only sterile release program. To that end we have developed a method for making transgenic L. cuprina using a piggyBac vector and an EGFP marker gene. We have also developed in Drosophila melanogaster a 2-component genetic system for controlling female viability. Females carrying both components of the system die unless fed a diet that contains tetracycline. We anticipate that the female-killing system will need to be optimised for L. cuprina in order to make a strain with the properties required for a male-only release program.     

Composition of the surface hydrocarbons from the vitelline membranes of dipteran embryos

Hydrocarbons were the major lipid class extracted by hexane from the vitelline membrane surface of dechorionated eggs of the house fly, Musca domestica, the New World screwworm, Cochliomyia hominivorax, the secondary screwworm, Cochliomyia macellaria, the green bottle fly, Phaenicia sericata, the sheep blow fly, Lucilia cuprina and the Mexican fruit fly, Anastrepha ludens. The length of time the embryos must be exposed to hexane with or without a small amount of alcohol in order to attain permeability was species-dependant. Long-chain n-alkanes comprised the major lipid class removed from vitelline membranes of all species except A. ludens where 2-methylalkanes were the major class. The range in size by the total number of carbon atoms in the hydrocarbons was: C23-C49 in C. hominivorax, C27-C33 in C. macellaria, C24-C35 in L. cuprina, C25-C36 in M. domestica, C25-C33 in P. sericata and C21-C51 in A. ludens. The major hydrocarbon component, expressed as percent of the total hydrocarbons, was n-nonacosane (C29) in C. hominivorax (40%), C. macellaria (43%), L. cuprina (38%), M. domestica (39%) and P. sericata (60%). However, in A. ludens, 2-methyloctacosane (32%) was the major hydrocarbon. Unsaturated hydrocarbons, monoenes (16%) and dienes (11%), were abundant only in A. ludens. Since prior studies indicated that the length of time the embryos must be exposed to hexane with or without a small amount of alcohol in order to attain permeability is species dependant, we suggest that the differences in hydrocarbon composition may contribute to this variation in lipid extractability.