Ecological analysis of field trials conducted to assess the potential of sex-linked translocation strains for genetic control of the Australian sheep blowfly, Lucilia cuprina (Wiedemann)

Field trials were conducted with translocation/eye colour (TE) strains of L. cuprina to measure the mating ability of the males under field conditions and assess their potential for suppressing sheep blowfly populations. Rates of increase in L. cuprina were highest in spring (3.9-9.1 per generation), consistently low during summer (0.1-0.6 per generation) and somewhat higher during autumn (1.1-3.4 per generation). The TE strains released had the potential to prevent population increases of this magnitude. Their failure to do so during these trials resulted from their low mating competitiveness (0.33) relative to that of field-reared males (1.0), inadequacy of the larval release method and the limited capacity of the experimental mass-rearing facility.     

Neuropeptidomics of the Australian sheep blowfly Lucilia cuprina (Wiedemann) and related Diptera

Insect neuropeptides are the most diverse and important group of messenger molecules that regulate almost all physiological processes, including behavior. In this study, we performed a combination of matrix assisted laser desorption ionization time of flight (MALDI-TOF) and electrospray ionization quadrupole time of flight (ESI-Q-TOF) mass spectrometry to analyze the peptidome of the brain and the neurohemal organs of the Australian sheep blowfly Lucilia cuprina and compared the data with those of related flies such as the gray flesh fly Sarcophaga (=Neobellieria) bullata; the cabbage root fly Delia radicum, the fruit fly Drosophila melanogaster, and the yellow fever mosquito, Aedes aegypti. Without counting low intensity signals of truncated peptides, 45 neuropeptides arising from 12 neuropeptide genes (adipokinetic hormone, CAPA-peptides, corazonin, extended FMRFamides, SIFamide, insect kinin, short neuropeptide F, NPLP-1 peptides, HUGIN-pyrokinin, sulfakinins, allatostatins A, putative eclosion hormone precursor peptide) were identified; sequences of extended FMRFamides were reported in a separate publication. The remarkable similarity of the peptidome of cyclorraphan flies, which contain a large number of ecologically important species, does not support the development of a species-specific neuropeptide-based insect pest control strategy. However, mass spectrometric approaches as shown here do not cover the entire peptidome or differences at the receptor level and it is possible that group-specific peptide ligands or receptors exist that escaped the detection.     

Genetic instability in mass-rearing colonies of a sex-linked translocation strain of Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae) during a field trial of genetic control

Summary Genetic breakdown occurred in a strain of Lucilia cuprina constructed for the purpose of genetic control of this pest. The strain incorporated autosomal recessive eye colour mutations linked in repulsion with a translocation involving the Y chromosome (male-determining) and two autosomes. In the original strain females had white eyes and males were wild type. The spontaneous breakdown involved a failure of the sex-limited inheritance of the eye colour mutations. Characteristically the frequency of white-eyed males increased rapidly in the strain, whereas the frequencies of the three other phenotypically recognizable breakdown products did not. This suggested that the white-eyed males had a selective advantage over both the wild type males and the other breakdown products. Genetic analysis revealed that recombination, which is normally rare in L. cuprina males, is considerably more frequent in the presence of a Y-autosome translocation, but that recombination alone was insufficient to account for the rate of increase of the white-eyed males in the colony. Genetic and cytological analysis of the breakdown products revealed that reversion of the multi-break translocation also occurred, and that many of the white-eyed males had either only a Y-single-autosome translocation or no translocation at all; thus these males were more fertile than the wild type multi-translocation males. In addition, under colony cage conditions the white-eyed males may have had a behavioural advantage in competition with the wild type males.     

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 .     

Xanthommatin biosynthesis in wild-type and mutant strains of the Australian sheep blowfly Lucilia cuprina

The synthesis of eye pigments has been studied in the seven eye color mutants of the Australian sheep blowfly, Lucilia cuprina. Six appear to be affected primarily in the synthesis of xanthommatin. In wild type, the onset of xanthommatin biosynthesis occurs midway through metamorphosis. Developmental patterns of accumulation of the xanthommatin precursors tryptophan, kynurenine, and 3-hydroxykynurenine have also been established for wild type. By determining the levels of these precursors in late pupae of the mutants, it has been shown that the mutant yellowish accumulates excess tryptophan and the mutant yellow accumulates excess kynurenine. The implications of these results—that yellowish lacks tryptophan oxygenase, thus failing to convert tryptophan to kynurenine, and that yellow lacks kynurenine hydroxylase (blocked in the conversion of kynurenine to 3-hydroxykynurenine)—have been confirmed. This has involved in vitro assays of tryphophan oxygenase and precursor feeding experiments. The precursor accumulation patterns are less clear for the other mutants.     

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.     

Aminopeptidases as potential targets for the control of the Australian sheep blowfly, Lucilia cuprina.

A series of experiments were carried out to investigate the role of proteinase enzymes in the growth of larvae of the sheep blowfly, Lucilia cuprina. First, instar larvae were incubated on an artificial growth media in the presence of various concentrations of inhibitors of all the major proteinase classes. Inhibitors of serine proteinases and aminopeptidases were found to cause significant growth inhibition and in some cases death of the larvae within 24 h, suggesting that these enzymes were the major classes involved in protein digestion in the gut of the insect. A second group of experiments analysed the effects of two inhibitors from the same or different proteinase classes in the growth media. Synergistic inhibition of larval growth was observed with the incorporation of inhibitors of serine proteinases and aminopeptidases. The results suggest that these classes of proteinases are both central to protein digestion in this insect, probably in the gut, and that the inhibition of both types of activity leads to an almost complete blockade of digestion. Testing in vivo gave similar results with infections on sheep skin inhibited by either serine proteinase or aminopeptidase enzyme inhibitors and the combination of both stopped the infection process. The role of aminopeptidases in larval metabolism and as potential targets for blowfly control agents is examined.     

Field trial of a compound chromosome strain for genetic control of the sheep blowfly Lucilia cuprina

Summary In 1979–80 a field trial of a compound chromosome (CC) strain of the Australian sheep blowfly Lucilia cuprina was conducted in the isolated Brindabella Valley, N.S.W. New genetic material was introduced into the strain before release by inducing 104 new CC elements by irradiation of recently captured field strains, and combining the resulting strains. Weekly releases, averaging 1.1 million larvae per week, were begun in November 1979 and continued to May 1980. Field-inseminated females were trapped weekly and their genotypes and those of their mates were determined through genetic testing. The proportion of wild X wild matings declined from 16% in December 1979 to 1% in April 1980. During this period the proportion of CC X CC matings rose from 50% to 90%. Larvae sampled from infested sheep had compound chromosomes, indicating that compound chromosomebearing females can successfully oviposit in the field. Trapping of flies resumed at the start of the 1980–81 season, without further releases. Progeny tests revealed the presence of both CC and wild flies. The proportions of CC X CC matings among field-inseminated females were 90% in October, 44% in November, nil in December, and 12% in January. No CC X CC matings were detected in 33 field-inseminated females trapped and tested during April, and 70 tested males reared from myiasis samples in April 1981 proved to be wild type. These results indicate that the CC strain overwintered in the field and strongly suggest that it bred in the field for at least one generation following the spring emergence before being eliminated from the population.     

The genetic bases of high-level resistance to diflubenzuron and low-level resistance to cyromazine in a field strain of the Australian sheep blowfly, Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae)

Abstract  Several genes on chromosomes IV and VI have a significant influence on high-level resistance to diflubenzuron in a strain of the Australian sheep blowfly from Tara, Queensland. Low-level resistance to cyromazine in the same strain is due to genes on these chromosomes with a gene (gene complex) in the sv marker region of chromosome IV being particularly important. For both insecticides, genetic background influences resistance status. If the results of the Tara strain prove typical for those of other populations, resistance to diflubenzuron in the Australian sheep blowfly has potentially significant consequences for woolgrowers.     

Innervation of the cercal sensilla on the ovipositor of the Australian sheep blowfly (Lucilia cuprina)

ABSTRACT. The ovipositor of the female sheep blowfly, Lucilia cuprina (Wied.) (Diptera: Calliphoridae), has a complement of cercal sensilla that includes long, medium and short tactile hairs, two campaniform domes, four olfactory pegs, and ten double-channelled gustatory hairs. This sensory array is suited to assess oviposition site resources, prior to and during the laying of an egg batch.
The tactile hairs and campaniform sensilla are each innervated by a single, tubular body containing dendrite. The olfactory pegs are each innervated by a single, moderately branched dendrite, which gains access to the external environment via pores at the bottom of deep grooves in the peg wall. The gustatory hairs fall into two categories. Four hairs have a single, tubular body containing dendrite at their base, and four unbranched dendrites running up to the hair tip which has a terminal pore. Six of the taste hairs have no tubular body containing dendrite at the base, and three unbranched dendrites running up to a terminal pore.     

Genetic architecture and adaptation: quantitative analysis of sheep and refuse tip populations of the Australian sheep blowfly, Lucilia cuprina

Phenotypic differentiation between geographic areas and between sheep and adjacent refuse tip populations was assessed by quantitative analysis of population samples of L. cuprina from New South Wales (Lismore) and Victoria (Mansfield). In addition the genetic structure of populations has been defined and compared by biometrical analysis techniques. For all morphological and fitness characters examined significant phenotypic differentiation was observed both between geographic localities and between sheep and non-sheep populations of each locality. Diallel analysis of the populations revealed architectural differences between sheep and non-sheep populations for both fecundity and egg hatchability. Sheep populations only, regardless of locality, displayed dominant gene effects on these fitness traits. The results suggest that refuse tip populations may be other than transients and that the differentiation may reflect differing patterns of adaptation and history of selection of the populations. The relevance of such differentiation to the successful establishment of a chemical and/or autocidal control zone is considered.     

The use of bridging systems to increase genetic variability in compound chromosome strains for genetic control of Lucilia cuprina (Wiedemann)

Summary Genetic variability in the non-compound portion of the genomes of compound-chromosome (CC) strains intended for genetic control can be increased by the use of bridging strains which can be crossed to both CC and normal strains. Two bridging systems are described for chromosome-5 CC strains of Lucilia cuprina. The first system relies on the established viability and fertility of males trisomic for chromosome 5R. Males carrying the (5L.YL)23 half-translocation, a C(5R), and a normal chromosome 5 were crossed successfully to a CC strain and a normal strain. The second system uses a pair of reciprocal whole-arm 4;5 translocations to generate gametes disomic for 5R and nullosomic for 5L, which in combination with C(5L)-bearing gametes form viable near-euploid offspring with only small duplications and deficiencies. These offspring (C(5L); (4L.5R)357; (4R.5R)194; (4L.4R)) were crossed successfully with both CC and T(4;5)357/ + individuals. The latter were in turn crossed successfully with normal strains. The T(Y;5)23 system allows replacement of the non-CC genome with wild material more rapidly than the T(4;5)357/T(4;5)194 system, but unlike the latter does not allow replacement of the Y chromosome in the CC strain. The double translocation system is currently being used in L. cuprina.     

Cuticular lipids of adults and puparia of the Australian sheep blowfly Lucilia cuprina (Wied.)

The presence of a strong contact component in the sex and ovipositing behavior of the sheep blowfly Lucilia cuprina Wied. prompted an investigation into the chemical composition of the cuticular wax of the adult male and female flies as well as that of the blowfly puparia. Thin-layer chromatography indicated that the lipids in all the waxes examined comprise hydrocarbons, nonglyceryl esters, triglycerides, free fatty acids, and hydroxy compounds, probably diglycerides and monoglycerides. Phospholipids were not detected. Straight-and branched-chain saturated compounds, the latter often pre-dominating, are present in the hydrocarbon, free fatty acid, and ester fractions. Unsaturated molecules were absent. The hydrocarbons resemble those of the cricket to some extent, but the absence of unsaturated compounds is in striking contrast to both the cricket and the cockroach. Pheromones may be present in the low molecular weight fatty acids obtained on brief extraction of the insects.     

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.     

Polytene chromosomes and linkage group-chromosome correlations in the Australian sheep blowfly Lucilia cuprina (Diptera: Calliphoridae)

The trichogen cell polytene chromosome maps for the Australian sheep blowfly Lucilia cuprina dorsalis R.-D. are presented. Correlations between the autosomal linkage groups and the polytene and mitotic chromosomes were accomplished using autosome-autosome and sex chromosome-autosome translocations. It is suggested that the sex chromosomes are largely inert.This work was completed during the tenure of a Fulbright fellowship and was supported in part by Public Health Service grant 2T1 GM 373-08.