Juvenile hormone synthesis by ring glands of the blowfly Lucilia cuprina

The major radiolabelled product released from ring gland and brain-ring gland complexes of third instar larval and pre-pupal stages of the sheep blowfly Lucilia cuprina upon incubation with L-[methyl-³H]methionine corresponded to one diastereomer of juvenile hormone III bisepoxide (JHB₃). Endocrine glands incubated with the juvenile hormone precursor 2E,6E-farnesoic acid released increased quantities of JHB₃, together with significant amounts of juvenile hormone III but not the isomeric methyl 2E-6,7-epoxyfarnesoate. Synthesis of JHB₃ was developmentally and neurally regulated. Ring glands and brain-ring gland complexes from third instar larvae released more JHB₃ than comparable preparations from pre-pupae, while isolated corpus allatum segments of the gland were more active than intact brain-gland complexes. These results reinforce the emerging status of JHB₃ as the characteristic juvenile hormone of dipteran insects. Arch. Insect Biochem. Physiol. 34:239–253, 1997. © 1997 Wiley-Liss, Inc.     

Hypersensitivity responses and repeated infections with Lucilia cuprina, the sheep blowfly.

, , and 1992. Hypersensitivity responses and repeated infections with Lucilia cuprina, the sheep blowfly. International Journal for Parasitology 22: 1175–1177. Sheep repeatedly infected with L. cuprina at 2- but not 4-week intervals developed partial resistance to infection after five infections, as measured by larval recovery. However, resistance did not persist for more than three infections. Skin weal responses were measured after injection of larval products simultaneously with each infection. The only correlation between weal size and larval recoveries occurred at infection 1 and indicated a relationship between skin sensitivity and innate rather than acquired resistance. The results suggest that resistance to L. cuprina can develop after repeated infections but that it is short lived and requires frequent larval exposure. A role for hypersensitivity responses was not confirmed by the weal responses but was suggested by the size of wound developed per larva recovered.     

Responsiveness to light of the circadian clock controlling eclosion in the blowfly, Lucilia cuprina

ABSTRACT. Eclosion in Lucilia cuprina (Wiedemann) occurs near dawn. The rhythm of eclosion persists in both darkness and constant light of high intensity (490μW cm^-2) with a period close to 24h. The sensitivity to light of the circadian clock controlling eclosion varies greatly according to the stage of the life cycle. During larval life the free running rhythm in darkness can be phase shifted by light pulses of 100μW cm^-2 intensity, with the transition from a Type 1 phase response curve to a Type 0, occurring with pulses of between 1 and 8h. Extending the last light period of LD to 24 h followed by constant darkness resets the phase of the rhythm by 12h, a transition from constant light to constant darkness initiates rhythmicity in flies made arrhythmic by being reared from eggs collected from adults maintained in constant light. After pupariation, the rhythm is relatively insensitive to light. Rhythmicity is sometimes induced by a transition from constant light to constant darkness, but the phase of the rhythm is not shifted by extending the last light period of LD before entering constant darkness. Repeated LD cycles applied after pupariation initiate and entrain the rhythm.     

Uptake and fate of specific antibody in feeding larvae of the sheep blowfly,Lucilia cuprina

Abstract. The quantity of specific antibody ingested by larvae of Lucilia cuprina and its fate after ingestion were studied in larvae grown on sheep and on an artificial diet. Larvae grown to late first or early second instar on sheep vaccinated with horse myoglobin contained 66% less specific antibody detected by enzyme linked immunosorbent assay than larvae grown to a similar stage on an artificial diet containing 75% serum from the same sheep. A similar result was obtained when larvae were grown to mid-third instar. Larvae grown on sheep to first or second instar contained approximately the same quantity of specific antibody per unit weight of larvae as those grown to third instar. Larvae grown on diet to third instar contained 22% less specific antibody per unit weight than those grown to first or second instar. In larvae grown on diet to late third instar, ingested diet retained 91 ± 12% of its original specific antibody activity in the crop, 50 ± 11% in the anterior midgut, 8 ± 2% in the posterior midgut and 13 ± 6% in the hindgut. The mean concentration of total immunoglobulin detectable in the haemolymph of individual third instar larvae grown on diet was 1.7 ± 2.8 ug/ml. Assays of specific antibody in the haemolymph of similarly reared larvae indicated that all or most of this immunoglobulin remained functional. The implications of the quantities and distribution of ingested functional antibody found in feeding larvae of L.cuprina are discussed in relation to the possibility of vaccinating sheep against these larvae and the selection of likely internal targets as sources of potential protective antigens.     

Tryptic and chymotryptic proteases released by larvae of the blowfly, Lucilia cuprina

Proteases released by larvae of the sheep blowfly have been suggested to have a primary role in wound formation and larval nutrition. Assays were carried out on two larval products to analyse the substrate specificity of these proteases, their abundance and approximate molecular weights. Tryptic and chymotryptic activities were found in both products though there were more chymotrypsin-like enzymes in products from 48 h cultures (CESP) than in product collected direct from 48 h larvae (LESP). Sodium dodecyi sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) gels incubated with azocasein showed plaques of major enzyme activity at molecular weights of 20,000 and 26,000 in LESP and at 20,000 in CESP. SDS-PAGE gels, when reacted with peptide substrates showed tryptic activity at 20,000 and 26,000 in LESP, whereas CESP showed only chymotryptic activity at 20,000 and higher molecular weights. The results suggest at least three enzymes, a trypsin and chymotrypsin in LESP, a chymotrypsin in CESP and a tryptic enzyme which is not stable to SDS-PAGE probably in both LESP and CESP. In addition, reactivity with elastase and plasmin substrates suggests the presence of enzymes with general effects on skin substrates and inflammatory pathways.     

Naturally occurring arrhythmicity in eclosion and activity in Lucilia cuprina: its genetic basis

ABSTRACT. Colonies of wild-type Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae) in our laboratory differed greatly in the degree of rhythmicity of their eclosion when exposed to light/dark cycles, constant darkness or constant light. Among colonies reared in the laboratory for thirty-four to eighty-eight generations and others for more than 360 generations, some were consistently rhythmic, others weakly rhythmic and others arrhythmic.
One colony with arrhythmic eclosion was studied further. Arrhythmicity of eclosion in the colony was found to result from homozygosis of a recessive allele (ary) on chromosome 5. Adults from this colony were arrhythmic in spontaneous flight activity under constant darkness though rhythmic under light/dark cycles.     

Monitoring and selection of resistance to pyrethroids in the Australian sheep blowfly, Lucilia cuprina

Field and laboratory populations of the Australian sheep blowfly, Lucilia cuprina (Wiedemann) (Calliphoridae), were surveyed by bioassay for possible resistance to the synthetic pyrethroids, a group of insecticides under development for blowfly control. A normal distribution of LC50 values was found using deltamethrin, the test pyrethroid, with no indication of specific resistance despite widespread use of deltamethrin on sheep to control the sheep body louse, Damalinia ovis (Schrank) (Trichodectidae). There was no cross-resistance to deltamethrin from existing organophosphate (OP) resistance nor from previous use of DDT. Selection with deltamethrin on a combined field strain, CSF85, increased the LC50 gradually over the first twenty generations until it stabilized at approximately 25x that of the unselected CSF85. This laboratory-induced resistance extended to other pyrethroids, cypermethrin (16x), cyhalothrin (25x) and cycloprothrin (10x), and increased the existing resistance of CFS85 to the OP diazinon (11x) and the carbamate, butacarb (83x).     

Resistance to two organophosphorus insecticides in New Zealand populations of the Australian sheep blowfly, Lucilia cuprina

Abstract. A treated-surface test was used in a bioassay to detect insecticide resistance to the organophosphorus compounds diazinon and propetamphos, in larvae of field strains of Lucilia cuprina in New Zealand during 1990/91 and 1991/92 (December-April). Mean LC₅₀s for both diazinon and propetamphos showed a significant increase between seasons although this is not necessarily indicative of a trend. Mean LC₅₀s for diazinon increased from 1.16 to 1.60 mg/1 (P< 0.001) and for propetamphos from 0.49 to 0.59 mg/1 (P< 0.005) in pooled populations. Diazinon and propetamphos resistance was significantly correlated in both seasons, suggesting side-resistance between the two insecticides.     

Regulation of Juvenile Hormone synthesis in the blowfly Lucilla cuprina

Abstract. Juvenile Hormone III bisepoxide synthesis by ring gland complexes from third-instar larvae of the blowfly Lucilia cuprina Weidemann (Diptera, Calliphoridae) was measured using a radiochemical assay in vitro. Hormone synthesis is regulated by three distinct mechanisms during development of the final larval instar up to the time of pupariation. The first type of regulation is detected as a rapid decline in hormone release coinciding with the final phase of commitment to pupariation. The second is a neurally mediated inhibition by the brain that acts at all stages of development in third-instar larvae. A protease-sensitive factor from brains of third-instar larvae causes dose-dependent reversible inhibition of Juvenile Hormone III bisepoxide synthesis. The third regulatory signal is a neural inhibition, observed in brain-ring gland complexes of prepupal stages. The first two levels of regulation appear to act early in the synthetic pathway for Juvenile Hormone (JH), whereas the third acts on the final steps of bisepoxide synthesis.     

Synthesis and deposition of chitin in larvae of the Australian sheep blowfly,Lucilia cuprina

Chitin synthesis in third-instar Lucilia cuprina larvae cultured at 23 °C was investigated using in vivo and in vitro systems, the latter with whole and with homogenized integuments. Synthesis was at a maximum between 24 and 48h after ecdysis from the second instar. Chitin was deposited in layers, and labeled GlcNAc was rapidly cleared from the hemolymph. In in vitro homogenate systems, the rapid conversion of UDP-([¹⁴C]GlcN)Ac to ([¹⁴C]GlcN)Ac and its 1-phosphate derivative contributed to the low incorporation of this precursor into chitin. The extent of the conversion was reduced by the addition of KCN or phenylthiourea. In in vivo and in vitro tissue systems the level of incorporation of ([¹⁴C]ClcN)Ac was higher than that of UDP-([¹⁴C]GlcN)Ac. However, in in vitro homogenate systems there was no difference unless UTP was added when the level of incorporation of only ([¹⁴C]GlcN)Ac was increased (by a factor of 9). Incorporation of UDP-([¹⁴C]GlcN)Ac, but not that of ([¹⁴C]GlcN)Ac, was decreased when larvae were deprived of food. Soluble oligosaccharides were detected in in vitro homogenate systems. They were formed during chitin synthesis and may represent newly initiated chitin chains. A reappraisal of current ideas on chitin synthesis in insects is needed.     

Effects of ovarian development and protein deprivation on the activity and locomotor responses of the blowfly, Lucilia sericata, to liver odour

Abstract. The spontaneous activity and locomotor responses to liver odour of females of the blowfly Lucilia sericata Meigen (Diptera: Calliphoridae) were examined in the laboratory using a rocking-box olfactometer. Spontaneous activity levels depended on fly age, previous exposure to protein and recent oviposition history; 1-day-old females, 3-day-old protein-fed females, and females that had recently oviposited exhibited lower levels of spontaneous activity than protein-deprived or fully gravid individuals. Changes in locomotor activity in response to liver odour also depended on previous exposure to protein. In protein-fed flies, activity decreased in response to liver odour and then increased on termination of an odour pulse. This response became more pronounced with age, broadly in line with the expected state of oocyte maturation. This pattern of change was not evident in protein-deprived females. The results are discussed in relation to the relative importance of protein and oviposition-site resources to L. sericata and the resultant age and ovarian development-stage biases evident in field catches.     

Visual and olfactory cue interaction in resource-location by the blowfly, Lucilia sericata

Abstract. The interactions between olfactory and visual cues in the landing responses of the blowfly Lucilia sericata (Meigen) (Diptera: Calliphoridae) were examined in a laboratory wind tunnel. The odour of liver and sodium sulphide solution released at 1 L/min, resulted in a greater number of landings, closer to, but downwind of, the central odour release point, than when odour was absent. Three-day-old protein-deprived females landed in greater numbers than protein-fed female flies of the same age; no difference existed between 6-day-old protein-fed or protein-deprived females. Six-day-old, protein-fed flies landed closer to the odour source than did 3-day-old, protein-fed flies. In the presence of odour, flies landed closer to the central release point when it was accompanied by a visual cue. No such effect of the visual cue was evident in the absence of odour. When a plain white sticky-surface (25 × 25 cm) was paired with an identical white surface plus odour, approximately equal numbers of flies landed when the surfaces were placed together; increasingly higher numbers of flies landed on the surface with the odour cue when the distance between the surfaces was increased. When a white surface with a visual cue was presented with the plain white surface plus odour, more flies landed on the white surface with the visual cue than on the plain white surface with odour when they were close together. However, as these two surfaces were moved apart, flies landed increasingly more on the surface with the odour than the surface with the visual cue. The results show that while odour cues may enhance the induction of landing by female L. sericata , visual cues are important when selecting a final landing site.     

The patterning of compensatory sugar feeding in the Australian sheep blowfly

Abstract. The pattern of feeding is described for males of the Australian sheep blowfly, Lucilia cuprina , with ad libitum access to either 0.1 M or 1.0 M glucose solution. Flies given the 0.1 M solution ingested nearly 3 times the volume taken by flies given the 1.0 M solution by eating meals of, on average, twice the size about 1.5 times as frequently. Flies were usually relatively inactive following a meal, with the extent of this post-prandial quiescence being a function both of meal size and concentration of sugar. Quiescence lasted only about 20% of the average intermeal interval, however, and there was no correlation between meal size and time to the next meal. The crop emptied more slowly when it contained 1.0 M rather than 0.1 M glucose solution and the crop was, on average, fuller at the beginning of a meal on the higher concentration. The volume of solution imbibed during a meal was positively correlated with time since the end of the preceding meal. The average crop volume at the end of a meal was similar in flies feeding on 0.1 M and 1.0 M solutions. The results are considered in relation to published information on control of feeding and compensation in the blowfly Phormia regina.     

Quantitative relationships between the ingestion of protein-rich material and ovarian development in the Australian sheep blowfly,Lucilia cuprina (Wied.)

The mature ovaries accounted for about 25% of dry weight and 30% of the protein content of wild-type (anautogenous) females of L. cuprina which had fed ad lib. on liver. The protein content of gravid females was 50% greater than at emergence. Protein ingested during the adult stage, therefore, plays an important role in ovarian maturation. The protein content of the ovaries of females fed measured amounts of liver exudate was from 37 to 52% of the amount of protein ingested.

Only limited ovarian development occurred in females fed only protein (high MW fraction of liver exudate or bovine serum albumin). The presence, in addition, of low MW components (low MW fraction of liver exudate or a salt mixture) was necessary for ovarian maturation. Quantitative feeding showed that the high and low MW fractions of liver exudate were, respectively, superior to bovine serum albumin and the salt mixture, which was based on the cation analysis of the low MW fraction of the exudate.     

Frequency‐dependent and density‐dependent larval competition between life‐history strains of a fly,Lucilia cuprina

1. Competition was created between the larvae of two life‐history strains of the blowfly Lucilia cuprina (Wiedemann) that have different requirements for larval resource acquisition. Adult females of one strain had the ability to mature eggs in the absence of adult feeding (autogeny) whereas the other strain lacked this ability. Autogeny shifts the burden of resource acquisition from adults to larvae, potentially leading to greater competition at this earlier life history stage. 2. A replacement series was used to determine the per‐capita competitive effect between strains relative to the intra‐strain effect, and density‐ and frequency‐dependent variation in this per‐capita effect was then evaluated. Evidence was found of competitive superiority of autogenous larvae when they occurred at a low frequency and low density, but their competitive ability was lost or reversed at higher frequencies and densities. 3. A dynamic competitive environment created by frequency and density dependence can account for the maintenance of genetic diversity for major life‐history traits. Such competition may explain why autogeny is rare in field populations of L. cuprina even although underlying genetic variation for the trait seems to be present.