Myosin light chain-2 mutation affects flight, wing beat frequency, and indirect flight muscle contraction kinetics in Drosophila.

We have used a combination of classical genetic, molecular genetic, histological, biochemical, and biophysical techniques to identify and characterize a null mutation of the myosin light chain-2 (MLC-2) locus of Drosophila melanogaster. Mlc2E38 is a null mutation of the MLC-2 gene resulting from a nonsense mutation at the tenth codon position. Mlc2E38 confers dominant flightless behavior that is associated with reduced wing beat frequency. Mlc2E38 heterozygotes exhibit a 50% reduction of MLC-2 mRNA concentration in adult thoracic musculature, which results in a commensurate reduction of MLC-2 protein in the indirect flight muscles. Indirect flight muscle myofibrils from Mlc2E38 heterozygotes are aberrant, exhibiting myofilaments in disarray at the periphery. Calcium-activated Triton X-100-treated single fiber segments exhibit slower contraction kinetics than wild type. Introduction of a transformed copy of the wild type MLC-2 gene rescues the dominant flightless behavior of Mlc2E38 heterozygotes. Wing beat frequency and single fiber contraction kinetics of a representative rescued line are not significantly different from those of wild type. Together, these results indicate that wild type MLC-2 stoichiometry is required for normal indirect flight muscle assembly and function. Furthermore, these results suggest that the reduced wing beat frequency and possibly the flightless behavior conferred by Mlc2E38 is due in part to slower contraction kinetics of sarcomeric regions devoid or partly deficient in MLC-2.     

Fourier analysis of wing beat signals: assessing the effects of genetic alterations of flight muscle structure in Diptera.

A method for determining and analyzing the wing beat frequency in Diptera is presented. This method uses an optical tachometer to measure Diptera wing movement during flight. The resulting signal from the optical measurement is analyzed using a Fast Fourier Transform (FFT) technique, and the dominant frequency peak in the Fourier spectrum is selected as the wing beat frequency. Also described is a method for determining quantitatively the degree of variability of the wing beat frequency about the dominant frequency. This method is based on determination of a quantity called the Hindex, which is derived using data from the FFT analysis. Calculation of the H index allows computer-based selection of the most suitable segment of recorded data for determination of the representative wing beat frequency. Experimental data suggest that the H index can also prove useful in examining wing beat frequency variability in Diptera whose flight muscle structure has been genetically altered. Examples from Drosophila indirect flight muscle studies as well as examples of artificial data are presented to illustrate the method. This method fulfills a need for a standardized method for determining wing beat frequencies and examining wing beat frequency variability in insects whose flight muscles have been altered by protein engineering methods.     

Rapid effects of progesterone on ciliary beat frequency in the mouse fallopian tube

Background  

The physiological regulation of ciliary beat frequency (CBF) within the fallopian tube is important for controlling the transport of gametes and the fertilized ovum. Progesterone influences gamete transport in the fallopian tube of several mammalian species. In fallopian tubes isolated from cows, treatment with 20 micromolar progesterone caused a rapid reduction of the tubal CBF. The aims of this study were to establish methodology for studying fallopian tube CBF in the mouse, as it is an important model species, and to investigate if progesterone rapidly affects the CBF of mice at nM concentrations.     

The effects of injection of amphetamine on female insemination in the black blow fly, Phormia regina (Diptera: Calliphoridae)

Abstract. An earlier study on the blowfly Phormia regina (Meigen) demonstrated that the injection of amphetamine (12 μg) depletes biogenic amine levels in the CNS. In the present study, P. regina females were injected with amphetamine (12 μg), each female was placed with three males and insemination success was determined. Amphetamine inhibited female insemination by 43.3% at 2–90 min post-injection and by 70% at 10–60 min post-injection. At 180–270 min post-injection, there was no significant inhibition of female insemination. This study indicates a possible role in insects for the biogenic amines in female insemination.     

Effects of temperature and temperature-steps on circadian locomotor rhythmicity in the blow fly Calliphora vicina

The free-running period (in darkness) of the locomotor activity rhythm in adult blow flies (Calliphora vicina) was temperature-compensated between 15 and 25 degrees C, showing Q(10) values between 0.98 and 1.04. Single steps-up (20 to 25 degrees C) or steps-down (20 to 15 degrees C) in temperature caused stable phase shifts of the activity rhythm, giving rise to temperature-step phase response curves (PRCs) with both advances and delays. Phase advances, however, were dominant for steps-up, and phase delays for steps-down; the two PRCs were almost "mirror images" of each other. Following protocols introduced by Zimmerman et al. [(1968) Temperature compensation of the circadian oscillation in Drosophila pseudoobscura and its entrainment by temperature cycles, Journal of Insect Physiology, 14, 669-684] for the rhythm of pupal eclosion in Drosophila pseudoobscura, the steps-up and steps-down PRCs for C. vicina were used to compute a theoretical PRC for a 6 h low temperature pulse, and from this a theoretical steady-state phase relationship of the locomotor activity rhythm to a train of such pulses making up a temperature cycle (18 h at 20 degrees and 6 h at 15 degrees C).     

Genotypic and environmental effects on flight activity and oviposition in the Glanville fritillary butterfly

Adverse environmental conditions constrain active flight and thereby limit reproduction in most insects. Butterflies have evolved various adaptations in order to thermoregulate, allowing females to search for nectar and to oviposit under unfavorable thermal conditions. We studied experimentally and with observational data the effect of low ambient temperatures experienced in the morning on the timing of oviposition and clutch size in the Glanville fritillary butterfly (Melitaea cinxia). Comparisons were made between individuals with different forms of the gene Pgi, encoding the glycolytic enzyme phosphoglucose isomerase, since naturally segregating variation at Pgi is known to be correlated with flight metabolic rate, flight performance, and fecundity. Experiencing low temperature in the morning delayed the initiation of oviposition and decreased clutch size. We used a thermal image camera to measure the thoracic surface temperature of butterflies immediately after voluntary flight. Single nucleotide polymorphism at Pgi was associated with thoracic temperature at low ambient temperatures. This has consequences for reproduction because females that are able to fly at lower ambient temperatures generally initiate oviposition earlier in the afternoon, when the environmental conditions are most favorable and the average egg clutch size is generally largest. These results suggest that variation in physiological and molecular capacity to sustain active flight at low ambient temperature has significant fitness-related consequences in insects.     

Sexual dimorphism in the pyrgomorphid grasshopper Phymateus morbillosus: from wing morphometry and flight behaviour to flight physiology and endocrinology

Abstract In the field, adult males of the grasshopper Phymateus morbillosus are able to fly for up to 1 min and cover up to c. 100 m, whereas females, although fully winged, are apparently unable to get airborne. Morphometric data indicate that the males are lighter, have longer wings, a higher ratio of flight muscles to body mass, and a lower wing load value than females. It was investigated whether this inability of females to fly is related to fuel storage, flight muscle enzymatic design and/or the presence and quantitative capacity of the endocrine system to mobilize fuels. In both sexes, readily available potential energy substrates are present in the haemolymph in similar concentrations, and the amount of glycogen in flight muscles and fat bodies does not differ significantly between males and females. Mass-specific activities of the enzymes GAPDH (glycolysis), HOAD (fatty acid oxidation) and MDH (citric acid cycle) in flight muscles are significantly lower in females compared with males, and mitochondria are less abundant in the flight muscles of females. There is no significant difference between the ability of the two sexes to oxidize various important substrates. Both sexes contain three adipokinetic peptides in their corpora cardiaca; the amount of each peptide in female grasshoppers is higher than in males.
Thus, despite some differences listed above, both sexes appear to have sufficient substrates and the necessary endocrine complement to engage in flight. It seems more likely, from the morphometric data above, that the chief reason for flightlessness is that P. morbillosus females cannot produce sufficient lift for flight; alternatively, the neuronal functioning associated with the flight muscles may be impaired in females.     

The redder the better: wing color predicts flight performance in monarch butterflies

The distinctive orange and black wings of monarchs (Danaus plexippus) have long been known to advertise their bitter taste and toxicity to potential predators. Recent work also showed that both the orange and black coloration of this species can vary in response to individual-level and environmental factors. Here we examine the relationship between wing color and flight performance in captive-reared monarchs using a tethered flight mill apparatus to quantify butterfly flight speed, duration and distance. In three different experiments (totaling 121 individuals) we used image analysis to measure body size and four wing traits among newly-emerged butterflies prior to flight trials: wing area, aspect ratio (length/width), melanism, and orange hue. Results showed that monarchs with darker orange (approaching red) wings flew longer distances than those with lighter orange wings in analyses that controlled for sex and other morphometric traits. This finding is consistent with past work showing that among wild monarchs, those sampled during the fall migration are darker in hue (redder) than non-migratory monarchs. Together, these results suggest that pigment deposition onto wing scales during metamorphosis could be linked with traits that influence flight, such as thorax muscle size, energy storage or metabolism. Our results reinforce an association between wing color and flight performance in insects that is suggested by past studies of wing melansim and seasonal polyphenism, and provide an important starting point for work focused on mechanistic links between insect movement and color.     

Assessing the effects of haematophagous ectoparasites on the health of nestling birds: haematocrit vs haemoglobin levels in House Wrens parasitized by blow fly larvae

We compared haematocrits and blood haemoglobin concentrations of nestling House Wrens Troglodytes aedon in nests with relatively large numbers of haematophagous larvae of the blow fly Protocalliphora parorum (>6.4 large larvae/nestling) and nests in which exposure to larvae was severely reduced or eliminated (0–2.0 larvae/nestling). Heavily parasitized nestlings showed no reduction in haematocrit levels but a significant 28% reduction in haemoglobin levels. Previous research on avian response to chronic blood loss suggests that parasitized nestlings may show normal volumes of red blood cells as a result of a rapid production of new cells in response to blood loss and, additionally, as a result of swelling of individual cells left in the bloodstream after parasite feeding. Rapid erythropoiesis places high numbers of haemoglobin-poor, immature cells into the bloodstream which would explain the reduced haemoglobin levels of parasitized nestlings. Our results refute the conclusion that larval feeding has little direct effect on nestling wrens in this parasite–host system, drawn earlier by Johnson and Albrecht (1993) based on examination of haematocrits alone. Our results also strongly suggest that researchers wanting to measure effects of blood-feeding parasites on birds measure haemoglobin levels, not haematocrits.     

Exploring the influence of individual courtship behaviors on male mating success in a blow fly

Male courtship behavior and displays influence female mating decisions, and therefore affect mating success in a diverse range of organisms. While there is substantial evidence confirming that females prefer males who invest more in courtship, less attention has been paid to the relative importance of individual behaviors, and the discrete sequences of courtship that result in mating success. The small hairy maggot blow fly, Chrysomya varipes, performs stereotyped courtship behaviors, involving orienting, tapping, waving, arching, wing vibration and mounting. This study aimed to quantify male investment in specific courtship behaviors, and compare courtship investment and behavioral transitions between males who gained mating success (successful males) and those that did not (unsuccessful males). Our results show that mating success was influenced by the behaviors orienting, tapping, arching and mounting. Behavioral transitions revealed a distinct pattern of behaviors leading to a mating attempt, and some differences were observed between successful and unsuccessful males. Overall, our findings suggest that female mating decisions were based on differences in specific male courtship behaviors. This detailed observational study has quantified multiple courtship behaviors for the first time in C. varipes, and highlights the importance of considering multiple behavioral traits when exploring the influence of male courtship on mating success.     

Growth modulating and cytotoxic effects of the peptide from hemolymph of blow fly Calliphora vicina (Diptera, Calliphoridae) in vitro

Biological activity of alloferon (AF), peptide, consisting of 13 a. a. isolated from hemolymph of experimentally infected blow fly Calliphora vicina was studied. AF in concentrations form 1 x 10(-5) to 250 microg/ml was added into the culture medium of the target cell lines K562, J-96, P388DI, Hep22a and 3T3B-SV40. First two days the peptide in concentrations 1 x 10(-5) and 1 x 10(-3) microg/ml in most cases stimulated the cell proliferative activity and suppressed the cell growth when applied in concentrations 10 and 100 microg/ml. Trend in growth modulating effect was dependent on duration of AF treatment. The peptide did not expressed cytotoxic effect with the exception of destruction of P388D1 cells that was registered after 96 h incubation in the medium initially contained 100 microg/ml AF. Simultaneously, cytotoxic and growth modulating effects of doxorubicin and cytosinarabinoside, as well as hybrid molecules, AF--cytosinarabinoside (cytal) and AF--doxorubicin (doxal) have been studied. Doxorubicin and cytosinarabinoside expressed greater growth inhibition effect compared to the hybrid molecules and AF itself. The results obtained with mass cell cultures were supported by experiments where P388D1 cells clonogenic capacity was tested. Besides, it has been demonstrated that AF rapidly penetrates into cytoplasm of J-96 cells and concentrates mainly into and around the cell nuclei.     

Free flight capacity determination in a sustained flight tunnel: effects of age and sexual state on the flight duration of Prostephanus truncatus

Abstract. . Adult Prostephanus truncatus (Horn) (Coleoptera: Bostrichidae) steering anemotactically upwind to a pheromone source in a wind tunnel, respond normally to, and alter their ground speed according to the direction and speed of, the movement of ground patterns beneath them. By manipulating this optomotor reaction component of their behaviour, studies were conducted on their flight duration. The results show a great deal of individual variation in flight duration, and the general data on single flights were skewed towards short flights. Pre-reproductive and inter-reproductive male and female beetles flew significantly longer than older beetles that had passed their peak of reproduction. There was no significant difference in the flight duration of male and female beetles at any age. The overall results suggest mat younger P. truncatus are capable of long-distance flights, and support the proposition that flight could be important in the spread of the beetle.     

Postnatal development in the big-footed bat,Myotis macrodactylus: wing morphology,echolocation calls,and flight

We studied the postnatal development of wing morphology and echolocation calls during flight in a free-ranging population of the big-footed bat, Myotis macrodactylus, using the mark-recapture methodology. Young bats were reluctant to move until 7 days of age and started fluttering at a mean age of 10 days. The wingspan and wing area of pups followed a linear pattern of growth until 22 days of age, by which time the young bats exhibited flapping flight, with mean growth rates of 0.62 mm/day and 3.15 mm²/day, for wingspan and area, respectively, after which growth rates decreased. Pups achieved sustained flight at 40 days of age. Of the three nonlinear growth models (logistic, Gompertz, and von Bertalanffy), the logistic equation provided the best fit to the empirical curves for wingspan and wing area. Neonates emitted long echolocation calls with multiple harmonics. The duration of calls decreased significantly between flutter (19 days) and flight (22 days) stages. The peak and start frequency of calls increased significantly over the 3-week period of development, but the terminal frequency did not change significantly over the development period.     

Maternal temperature has different effects on the photoperiodic response and duration of larval diapause in blow fly ( Calliphora vicina ) strains collected at two latitudes

Abstract .The blow fly Calliphora vicina Robineau-Desvoidy (Diptera: Calliphoridae) has a wide distribution across northern and temperate Europe. It has a facultative, maternally-induced larval diapause in response to short days. The photoperiodic response, measured at 15 and 20°C, of two populations was compared. A southern population (originating at 51° N) was sensitive to temperature at all daylengths; the incidence of diapause was greatly reduced at 20°C compared with 15°C. The photoperiodic response of a northern population (from 65° N) was sensitive to temperature only in long days; in short days (< 14 h of light) the response of this strain was identical at each temperature.
Variation in parental photoperiod and temperature were found to affect the duration of larval diapause, indicating a role for maternal effects in diapause intensity as well as incidence. However, the between-strain variation was greater than that within strains, indicating qualitative differences in diapause response. These differences may arise from the ecological conditions at the points of origin of the two strains. The northern strain from the harsher climate has a more intense diapause that follows a relatively temperature-insensitive photoperiodic response. In contrast, the southern strain has a shallow diapause and its photoperiodic response may be overridden by the experience of concurrent high temperature.     

Nutrient absorption and utilization by wing and flight muscle morphs of the cricket Gryllus firmus: implications for the trade-off between flight capability and early reproduction

Absorption efficiency (AD, approximate digestibility, assimilation efficiency) of various macronutrients and conversion of absorbed nutrients to biomass (ECD) were compared among the two types of flightless morph and the flight-capable morph of the cricket, Gryllus firmus. No biologically significant phenotypic or genetic difference in AD for carbohydrate, protein or lipid was observed among morphs fed either a high-nutrient (100%) or a low-nutrient (25%) diet. Thus, previously-documented differences among adult morphs in carbohydrate and lipid content must be caused by processes other than variation in nutrient absorption by morphs during adulthood. Relative absorption efficiency of total dry mass of food by morphs of G. firmus appears to be a valid indicator of relative AD of total calories. Morphs did not differ phenotypically or genetically in the excretion of end products of nitrogen metabolism (uric acid, hypoxanthine plus xanthine) on either the high nutrient or the low nutrient diet. Nutritional indices corrected for excreted nitrogenous metabolites were very similar to uncorrected indices, and the pattern of variation among the morphs was the same for corrected or uncorrected values. Each of the two types of flightless morph converted a greater proportion of absorbed nutrients into body mass, mainly ovaries, and allocated a smaller proportion of assimilated nutrients to respiration than did the flight-capable morph. Moreover, the trade-off between respiration and early reproduction was substantially magnified on the low nutrient diet. These results extend previous findings of a trade-off between flight capability and early reproduction in wing-polymorphic Gryllus species (1) to diets of very different nutrient quantity, and (2) to flightlessness arising from different causes: blockage of flight muscle development in juveniles vs histolysis of fully-developed flight muscles in adults.     

Histochemical localization of trehalase activity in dorsal flight muscle of the flesh fly Sarcophaga bullata with light and electron microscopy.

Trehalase activity in flight muscle of the flesh fly Sacrophaga bullata is detected histochemically at light- and electron-microscopic levels by using diaminobenzidine, glucose oxidase and peroxidase in the incubation medium. The association of trehalase activity with the inner mitochondrial membrane is confirmed. Biochemical assay shows that about 50% of the initial total trehalase activity is lost from the tissue during the histochemical processing and about 50% remains for histochemical detection.