Cold-hardiness: a component of the diapause syndrome in pupae of the flesh flies, Sarcophaga crassipalpis and S. bullata

ABSTRACT. Diapausing pupae of Sarcophaga crassipalpis Macquart and S. bullata Parker reared at 20 or 25C readily survive exposure to - 10C for at least 25 days. In contrast, non-diapausing pupae produced by a variety of means are consistently intolerant of the low temperature. Non-diapausing pupae are not immediately killed by exposure to -10C: pupae exposed to the low temperature for up to 3 days proceed with pharate adult development but ultimately die before adult eclosion. Unlike many temperate zone insects, diapausing flesh fly pupae do not require a period of chilling for induction of cold-hardiness, and the attribute of cold-hardiness cannot be separated from other features of the diapause syndrome. Some cold-hardiness is already acquired during the third larval instar: diapause-destined larvae exposed to -10C are more successful in pupariating than non-diapause-destined larvae of the same age.     

The diffusional and osmotic water permeability of the hindgut cuticle in larval Sarcophaga bullata

The diffusional water permeability (P_D) and the pressure filtration coefficient (L_P) of isolated larval hindgut cuticle of the fleshfly, Sarcophaga bullata, were measured using tracer techniques coupled with a simple mathematical model system based on equations of non-equilibrium thermodynamics. Data obtained from the model system were matched to experimental tracer data by means of a mathematical optimization scheme. The following parameter values were obtained: P_D for tritiated water = 1.02 × 10^?6 cm-sec^?1, and L_P = 9.18 × 10^?11 cm³-dyn^?1-sec^?1. These results are now being used to determine reflection coefficients (σ's) and solute mobilities (ω's) for the cuticle system in an attempt to gain an understanding of the mechanisms controlling solute and water movements across the hindgut wall.     

Cytoskeletal F-actin patterns in whole-mounted larval and adult salivary glands of the fleshfly, Sarcophaga bullata.

The patterns of filamentous actin were analysed in different larval, pupal and adult stages in the salivary glands of the fleshfly Sarcophaga bullata. Using the rhodamine labelled phalloidin staining method in combination with detergent extraction specific actin filament distribution was detected. The salivary glands which are histolysed during the process of metamorphosis show distinct cellular morphology and actin filament patterns in larvae and adults. The large third instar larval salivary gland cells contain a well developed apicolateral microvillar zone. In third instar larvae this microvillar zone invaginates and expands in the basal part of the lateral membranes. Larval salivary gland cells also contain numerous parallel basal actin bundles. The larval glands are histolysed during metamorphosis and adult glands are formed out of the imaginal cell group. At the onset of metamorphosis these basal actin bundles form a network of crossing bundles. The filamentous actin patterns of the proximal part of adult gland cells is confined to the apicolateral microvillar membranes. The cells in the distal, tubular part of the adult salivary glands show intense staining of their folded lateral membranes.     

Stage specific larval,pupal, and adult cuticles in the tracheal system of Sarcophaga bullata

Successive tracheal cuticles of the dorsal longitudinal trunks are studied with the electron microscope. Minor differences seen at the light microscope level are seen as major qualitative and quantitative ones at the ultrastructural level. The larval and pupal cuticles are secreted by similar epithelial cells; these possess large polytene chromosomes. Cell division and possibly cell replacement occur prior to adult cuticle secretion. The findings are discussed in terms of cell specificity, intra- and inter-cellular pattern formation. This simple epithelium, the individual cells of which are capable of producing different cuticles, is interesting since the system is also shown to be responsive to hormone application.     

The photoperiodic clock in the flesh-fly, Sarcophaga argyrostoma

Larval cultures of the flesh-fly, Sarcophaga argyrostoma, were raised in experimental light cycles with periods (T) of 21 to 72 hr, each cycle containing a photoperiod of 4 to 20 hr of white light. This ‘resonance’ technique revealed periodic maxima (～24 hr apart) of pupal diapause, thereby demonstrating an endogenous circadian component in the photoperiodic clock. The positions of these maxima of pupal diapause suggested that the oscillation, like that controlling the pupal eclosion rhythm in Drosophila pseudoobscura, is ‘damped out’ by photoperiods longer than about 11 to 12 hr, but restarts at dusk whereupon it runs with circadian periodicity in a protracted dark period. With photoperiods shorter than 12 hr, however, the two diapause maxima were less than 24 hr apart, suggesting that an additional component, possibly a ‘dawn hour-glass’, was modifying the position of the first peak.Both photoperiod and the period of the driving light cycle (T) were shown to affect the length of larval development (the sensitive period) and the number of calendar days needed to raise the incidence of pupal diapause to 50 per cent (the required day number, RDN). Peaks of diapause induction were shown to be the result of an interaction between a long sensitive period (slow development) and a low RDN, whereas troughs in diapause induction were the result of an interaction between a short sensitive period (fast development) and a higher RDN.Larvae of S. argyrostoma are unable to distinguish (in a photoperiodic sense) between 12 and 18 hr of red light (600 nm).     

Mevalonate biosynthesis in the fly,Sarcophaga bullata

Radioactive mevalonic acid (MVA), the committed substrate in isoprenoid biosynthesis, is synthesized from [2-¹⁴C]acetate in the presence of various co-factors by enzyme fractions from non-axenic and axenic Sarcophaga bullata larval homogenates. This mevalonic acid, in the lactone form, has been rigorously characterized by several chromatographic and physical analyses. Since the axenic and non-axenic homogenate incubations were very similar, the role of microflora in this synthesis is not considered significant. Of several larvae tissues examined, gut was the most active in MVA synthesis and fat body the least. However, all tissues examined had some MVA synthetic activity. Non-radioactive MVA was isolated quantitatively from larvae, pupae, and adults, and thus it would appear that there is a pool of MVA available for isoprenoid biosynthesis throughout development.     

Neuromuscular junctions and L-glutamate-sensitive sites in the fleshfly, Sarcophaga bullata.

Sites have been located on retractor unguis and trochantal depressor muscle fibres of Sarcophaga which respond to iontophoretic application of l-glutamate. No such sites could be found on flight muscle fibres. Ultrastructural examination of the three muscles reveals differences between the muscles in the positions of the neuromuscular junctions. A correlation can be made between the sites of the neuromuscular junctions and the iontophoretically sensitive sites. The possibility of l-glutamate fulfilling a transmitter rôle in these muscles is discussed.     

Mapping of serotonin-immunoreactive neurons in the larval nervous system of the flies Calliphora erythrocephala and Sarcophaga bullata

Summary Serotonin-immunoreactive (5-HTi) neurons were mapped in the larval central nervous system (CNS) of the dipterous flies Calliphora erythrocephala and Sarcophaga bullata. Immunocytochemistry was performed on cryostat sections, paraffin sections, and on the entire CNS (whole mounts).The CNS of larvae displays 96–98 5-HTi cell bodies. The location of the cell bodies within the segmental cerebral and ventral ganglia is consistent among individuals. The pattern of immunoreactive fibers in tracts and within neuropil regions of the CNS was resolved in detail. Some 5-HTi neurons in the CNS possess axons that run through peripheral nerves (antenno-labro-frontal nerves).The suboesophagealand thoracico-abdominal ganglia of the adult blowflies were studied for a comparison with the larval ventral ganglia. In the thoracico-abdominal ganglia of adults the same number of 5-HTi cell bodies was found as in the larvae except in the metathoracic ganglion, which in the adult contains two cell bodies less than in the larva. The immunoreactive processes within the neuropil of the adult thoracico-abdominal ganglia form more elaborate patterns than those of the larvae, but the basic organization of major fiber tracts was similar in larval and adult ganglia. Some aspects of postembryonic development are discussed in relation to the transformation of the distribution of 5-HTi neurons and their processes into the adult pattern.     

A developmental study of the cuticular hydrocarbons of Sarcophaga bullata.

The accumulation of cuticular hydrocarbon was measured throughout the life of Sarcophaga bullata. Less than 5 μg hydrocarbon per insect are present until the third larval instar when synthesis increases the quantity present to 10 to 20 μg by pupariation. The rate of synthesis increases at this time to 5 to 8 μg/day and continues until 40 to 45 μg are present per insect. This amount remains constant until several days before the pupal-adult ecdysis when synthesis again occurs. The rate of synthesis by these pharate adults is >20 μg/day. When the adult emerges it contains between 90 and 100 μg which increases slightly during the adult stadium. Two periods of rapid accumulation of cuticular hydrocarbon are observed: (1) during pupariation and the 3 day period following pupariation, and (2) during the 4 day period preceding the pupal-adult ecdysis. When pupariation is inhibited by contact with water, the rate of hydrocarbon biosynthesis also fails to increase.     

Moulting hormone,juvenile hormone and the ultrastructure of the fat body of adult Sarcophaga bullata (Diptera)

Summary In the ovoviviparous fly, Sarcophaga bullata, vitellogenesis is cyclic; a process reflected in ultrastructural changes in the fat body cells and oenocytes. At eclosion the larval fat body has not yet completely disappeared. During vitellogenesis the fat body cells are specialized for intensive protein synthesis showing a very extensive RER and numerous invaginations of the plasma membrane. These features disappear when the eggs descend into the oviducts to complete embryogenesis. The predominant feature of the oenocytes is their very prominent SER. The fat body cells of the males are never as specialized for protein synthesis as those of the females. Feeding of ecdysterone to males for 3 or more days induces a rather extensive subcellular apparatus for protein synthesis, i.e., invaginations of the plasma membrane and an extensive RER. Juvenile hormone is completely ineffective in this respect. Both ecdysterone and juvenile hormone have pronounced but different effects on the oenocytes of males.     

Food conversion efficiency by fleshfly larvae, Sarcophaga bullata

ABSTRACT. The efficiency of the conversion of the digested food (ECD) (liver) during the growing period of the larva of Sarcophaga bullata was determined. The values for ECD in four different determinations were 67.8, 69.1, 64.1 and 63.4%. These values are of the same order as ECD determined for some phytophagous caterpillars under the best conditions of food utilization, and seem to represent the highest values obtainable in a multicellular organism. It is concluded that such high efficiency rates have as prerequisites: fast growth in the larvae; and in the food, easy availability, ingestibility, and digestibility, high nutritional value, and reasonably high water content. Bacterial decomposition of the food during the test period appeared scarcely to influence the apparent ECD.     

Allatostatin-like immunoreactivity in the optic lobe of the fly Sarcophaga bullata.

An antiserum against Diploptera allastostain 1 (Dip-AST1) was used to map the distribution of allatostain containing neurons in the optic lobes of the fly Saccrophaga bullata. Strongly immunoreacting neurons were found in two areas of the optic ganglia, namely, the medulla and the area between medulla and lobula. These cells were generally interneurons arborizing the base of the medulla. The positive reaction of specific populations of the optic lobe neurons against allatostain antiserum suggests some role for this neuropeptide in the visual physiology of the fly.     

Genomic organization in the flesh fly Sarcophaga bullata

The genome of the flesh fly Sarcophaga bullata has been characterized both cytologically and biochemically. S. bullata has a haploid DNA level of 0.61 picograms which is five times larger than the haploid genome size of Drosophila melanogaster. Reassociation kinetics of Sarcophaga DNA shows that its sequence organization is very similar to that of D. melanogaster in having a very large proportion of single copy DNA (81%) and only small amounts of highly and moderately repetitive DNA (9% and 6%, respectively). cRNAs from all three sequence classes were prepared and their cytological distributions on diploid and polytene cells determined by in situ hybridization. The cytological distribution of the highly repetitive probe was found to be restricted to the centromeric heterochromatin of two of the five autosomes and this sequence class was also found to be markedly underreplicated in polytene foot-pad cells. No highly repetitive DNA was localized on either of the sex chromosomes, but only on the two large centromeric regions of chromosomes C and E. Moderately repetitive DNA was found uniformly distributed on all of the autosomes in both testis and polytene foot-pad squashes. As in the case of the highly repetitive sequence probe, no moderately repetitive DNA was detected on either the X or Y chromosomes. Moderately repetitive DNA in Sarcophaga was also shown to have the Drosophila type pattern of sequence interspersion with a moderately repetitive element of 5,000 nucleotides adjacent to a unique element of greater than 10,000 nucleotides. The Sarcophaga genome is the largest for which this type of interspersion has so far been demonstrated.     

Entomophthora bullata as a pathogen of Sarcophaga aldrichi

Adults of Sarcophaga aldrichi killed by Entomophthora bullata were found in the field 7 days after emergence. Resting spores were not found until 6 days later, although conidia were formed from the first incidence of mortality. It is suggested that the life cycle of the fungus involves alternating generations of conidia and thick-walled overwintering resting spores.     

Mendelian inheritance of pupal diapause in the flesh fly, Sarcophaga bullata

Pupal diapause (dormancy) in the flesh fly, Sarcophaga bullata, is induced by short-day photoperiods and low temperature. In this study, the inheritance mode of diapause was investigated by crossing a nondiapausing (nd) strain of S. bullata with 2 diapausing strains having different diapause capacities. The results consistently indicated that diapause incidence is inherited in a simple Mendelian pattern, thus a single gene or a small gene cluster linked to the photoperiodic clock controls the seasonal response of diapause. The fact that the nd strain lacked daily rhythmicity in adult eclosion and showed altered expression of 2 circadian clock genes suggests that the photoperiodic and circadian clocks are related through a shared molecular component in S. bullata.