Purinoceptors in blood feeding behaviour in the stable fly, Stomoxys calcitrans

Abstract Both sexes of the stable fly, Stomoxys calcitrans L. (Diptera: Muscidae), have receptors on their mouthparts that mediate blood feeding. The potency ranking of the adenine nucleotides (ATP > ADP > AMP > adenosine) in eliciting feeding and suppressing the NaCl-sensitive cell may indicate the involvement of a P₂-type receptor. This is supported by the lack of effect on feeding by methyl xanthines. Feeding-related behavioural and electrophysiological results demonstrate that the potency of CH₃-S-ATP is not greater than that of ATP. These ATP-mediated responses are antagonized by ANAPP₃. Results support the conclusion that the putative 'ATP receptor' involved in stable fly phagostimulation resembles the P_2x purinoceptor of vertebrates.     

Circadian activity pattern in the stable fly, Stomoxys calcitrans

Abstract. Activity rhythms of Stomoxys calcitrans (L.) were assessed under constant conditions using actographs.In LD 12:12 h, activity was almost completely diurnal, had a 24 h period, and peaked 4 h after lights-on.In constant darkness, activity peaked in the early afternoon and the period of the rhythm was c. 26 h, indicating a circadian pacemaker.A delay in feeding shortened the free-running period in constant dark to c. 24 h.In constant light, flies were arrhythmic.Starvation also affected activity, with 24- and 48-h starved flies being as much as 20 times more active than recently fed flies.     

Electroantennogram responses of the stable fly, Stomoxys calcitrans, to components of host odour

Abstract. Electroantennograms (EAGs) were recorded from laboratory-reared male and female Stomoxys calcitrans (L.) in response to a range of synthetic chemicals known to be electrophysiologically-active for other biting flies. Of the eight compounds initially tested, only two - 1-octen-3-ol and 3-methylphenol - consistently elicited larger electroantennograms (EAGs) than did control treatments; 1-octen-3-ol was the most potent. EAG recovery time was inversely correlated with EAG amplitude. EAGs recorded with primary C₂-C₁₂ carbon chain-length primary aliphatic alcohols peaked at octan-1–ol with pentan-1-ol, hexan-1-ol and heptan-1-ol also eliciting EAG responses significantly larger than the controls. When different C₈ carbon chain compounds and nonane were tested: 1-octen-3-ol elicited the largest EAGs followed by, in decreasing activity, octan-1-ol, 1-bromooctane, octan-3-ol, octanal, 2-octanone, octanoic acid and nonane. The EAG response of 1-octen-3-ol increased sigmoidally with dose, with the threshold at between 2 and 20 ng, and the peak response at 200 μg on filter paper. EAGs larger than control were also elicited by entrained ox odour and ox breath. The behavioural implications are discussed.     

Characterization of mannosyl transferases during the pupal instar of Stomoxys calcitrans (L)

Particulate fractions (10,000g) from pupae of Stomoxys calcitrans transfer [¹⁴C]-mannose from GDP-[¹⁴C]-mannose to dolichol monophosphate and proteins. Production of the mannosyl lipid was inhibited by Mn²⁺, UDP, GMP, GDP, and EDTA. The insect growth regulator diflubenzuron had no effect on mannosyl transferase activity. Dolichol monophosphate and Mg²⁺ stimulated mannosyl transferase activity. The mannosyl lipid product was identified as mannosyl-phosphoryl-dolichol (Man-P-Dol). The apparent K_m and V_max values for the formation of Man-P-Dol using GDP-[¹⁴C]-Man while holding dolichol phosphate constant were 2.4 ± 0.9 μM and 9.4 ± 2.3 pmol Man-P-Dol·min^?1·mg^?1 protein, respectively. The apparent K_m and V_max values using dólichol phosphate while holding GDP-Man constant were 2.2 ± 1.2 μM and 18.5 ± 1.7 pmol Man-P-Dol·min^?1·mg^?1 protein.     

Absence of female-specific protein in the hemolymph of stable fly Stomoxys calcitrans (L.) (Diptera: Muscidae)

Changes, during the reproductive cycle, in fat body, hemolymph, and ovarian proteins of the stable fly Stomoxys calcitrans were characterized quantitatively and qualitatively using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Protein content of all three tissues increased after blood feeding. Fat body protein increased first, followed by hemolymph and ovarian proteins. SDS-PAGE failed to identify vitellogenin in both female hemolymph and fat body samples. No single protein or group of proteins predominated at any stage of the reproductive cycle. Comparisons between male and female stable fly hemolymph and fat body proteins failed to detect female-specific proteins. Female-specific proteins, however, were detected in the hemolymph of four other species of Diptera.     

Retention and attempted mechanical transmission of Ehrlichia risticii by Stomoxys calcitrans

Abstract. The ability of adult Stomoxys calcitrans (L.) (Diptera: Muscidae) to retain viable Ehrlichia risticii (Rickettsiaceae), the aetiologic agent of Potomac horse fever (PHF), and mechanically transmit the pathogen from citrated bovine blood artificially infected with E.risticii to susceptible mice was studied. Viable E. risticii were found in the digestive tract of S.calcitrans 3h after the flies had engorged to repletion on infected blood; however, no E. risticii were detected in flies ≥2 days after feeding. Subsequently, groups of S.calcitrans were fed for 20 s on infected blood, then fed to repletion on mice 30, 60,130, 180 or 220 min after having feeding interrupted. Mice displayed no clinical signs of PHF and did not produce anti- E. risticii antibodies when assayed 30 days after S. calcitrans had fed. Although S.calcitrans were able to harbour viable E.risticii for at least 3h, transmission of the disease agent to susceptible mice during interrupted feeding was not demonstrated under these experimental conditions.     

Stable fly, Stomoxys calcitrans, mouthpart removal influences stress and anticipatory responses in mice

Abstract Biting fly attack induces a variety of stress and anxiety related changes in the physiology and behaviour of the target animals. Significant reductions in pain, or more appropriately, nociceptive sensitivity (latency of a foot-lifting response to an aversive thermal stimulus), are evident in laboratory mice after a 1 h exposure to stable flies, Stomoxys calcitrans. The role of the various components of biting fly attack in the development of this stress-induced reduction in pain sensitivity (analgesia) is, however, unclear. This study demonstrates that fly-naive mice do not exhibit a stress-induced analgesia when exposed to stable flies whose biting mouthparts have been removed. In contrast, mice that have been previously exposed to intact stable flies exhibit significant analgesia when exposed to flies that are incapable of biting. However, the level of analgesia induced is lower than that elicited by exposure to intact stable flies. Exposure to non-biting house flies, Musca domestica , has no effect on nociceptive sensitivity. It appears that the actual bite of the stable fly is necessary for the induction of analgesia and probably other stress and anxiety associated responses in fly naive mice. However, mice rapidly learn to recognize biting flies and exhibit significant, possibly anticipatory analgesic responses to the mere presence of biting flies.     

Electroantennogram responses of the stable fly, Stomoxys calcitrans, to carbon dioxide and other odours

ABSTRACT. Electroantennograms (EAGs) were recorded from Stomoxys calcitrans (L.) in response to increasing concentrations of carbon dioxide in an airstream. The magnitude of the EAG increases logarithmically from +0.023% carbon dioxide up to approximately +2.0% where a maximum is reached. Flies deprived of food for 48–50 h are more responsive to small increases in the carbon dioxide concentration than those deprived of food for only 20–23 h. It is concluded that the sensitivity of carbon dioxide receptors on the antennae of S. calcitrans increases as hunger develops. EAGs were also recorded in response to cattle odour, odour from fresh cattle faeces, expired human breath, acetone, and l-octen-3-ol. Acetic acid vapour causes a reversal of the usual EAG response indicating inhibition.     

Characterization of proteolytic enzymes of the midgut and excreta of the biting fly Stomoxys calcitrans

Proteolytic enzymes were characterized in the midgut and the excreta of the stable fly Stomoxys calcitrans (L) with proteins, synthetic substrates, and inhibitors. Inhibition studies suggested trypsinlike activity in sugar-fed fly midguts, whereas excreta and blood-fed fly guts exhibited other proteases. Trypsinlike activity in midguts removed 20 and 30 h after a blood meal increased from 20% to 50% of the total proteolytic enzymes present. Trypsinlike activity was inhibited with human sera, trypsin-specific inhibitors, and a protein isolated from the stable fly thorax. When human albumin and globulin fractions were incubated with trypsinlike enzymes isolated from the midgut and excreta, the albumin fraction was less inhibitory than the globulin fractions and was readily hydrolyzed by the proteolytic enzymes. These results may indicate that the proteolytic enzymes produce an abortive complex with the globulin fractions of the sera. Such a complex may explain the temporary inhibition of proteolysis by the blood meal. Soybean trypsin inhibitor fed to stable flies caused 50% inhibition in proteolytic activity in the midguts of sugar-fed stable flies and 25% inhibition in the midguts of blood-fed stable flies. Complete inhibition of proteolytic enzyme activity was achieved only in vitro. pH profiles of proteolytic enzyme activity isolated from the excreta of blood-fed stable flies indicated that several proteolytic enzymes were excreted.     

Effects of carbon dioxide, acetone and 1-octen-3-ol on the flight responses of the stable fly, Stomoxys calcitrans, in a wind tunnel

Abstract. The flight behaviour of Stomoxys calcitrans (L.) in odour plumes containing carbon dioxide, acetone or l-octen-3-ol was assessed from video recordings. A downwind bias was evident in clean air, whereas all three test chemicals elicited upwind anemotaxis. Response thresholds were ∼0.006% for CO2, between 0.001 and 0.01 μg/l for acetone, and ∼0.0002 u.g/1 for l-octen-3-ol. Sinuosity (° cm^-1) and angular velocity (° s^-1) increased with C0₂ concentration, but velocity (cm s"¹) decreased. Similar, but less clear effects were observed for acetone and l-octen-3-ol.     

Isolation and in vitro synthesis of trypsin from the biting fly,Stomoxys calcitrans

The synthesis of [3H]trypsinlike enzyme by the fat body was followed in Stomoxys calcitransin vitro using a radioimmunoassay (RIA) developed against mammalian trypsin. Using high specific activity [3H]valine, trypsinlike activity was followed in midgut epithelial cells, thoracic muscle, and fat body removed from sugar-fed flies. Excreta protease of S. calcitrans was partially purified using charge and hydroxylapatite gel chromatography. Seventy-five percent of the enzyme eluted from these gels was inhibited by tosyl-L-lysine chloromethyl ketone HCI (TLCK) and was classified as trypsinlike. Electrophoresis of the trypsinlike enzyme indicated that it was only 50% pure. Trypsinlike activity from S. calcitrans bound to α₁-globulin IV-I and formed a complex that was dissociated on a P-100 Bio-Gel column. Binding between the protease and the α₁-gobulin IV-I caused a 1.4-fold increase in the apparent molecular weight of the protease on the P-100 Bio-Gel column. Trypsinlike activity was characterized in the midgut and excreta by affinity binding to covalently linked TLCK and tosyl-L-lysine chloromethyl ketone HCI (TAME)Sepharose 4B gels. Between 50% and 55% of the excreta protease and 5669% of the midgut protease bound to the affinity gels and was trypsinlike. Protease activity that did not bind to the gels was not inhibited by TLCK and did not have the esterolytic activity of trypsin.     

Effects of two blood-feeding regimes on mortality and female reproduction in a laboratory colony of stable flies,Stomoxys calcitrans

Abstract. Stable flies (Stomoxys calcitrans L.) deprived of a bloodmeal until 3 days post-emergence had higher mortality rates than control flies fed from the day of emergence. Fat bodies of deprived females required one more bloodmeal to reach maximum size, and maximum size was smaller, than fat bodies of control females. Ovarian development did not commence prior to feeding in deprived flies, and proceeded more slowly thereafter, resulting in a one bloodmeal delay in egg maturation in deprived flies. Deprived females produced fewer (54.7, SD 2.8) eggs than controls (75.9, SD 3.7) and eggs from deprived females were smaller (mean length 684.0 μrn) than control females' eggs (mean length 1165.7 μm).     

Observations on the mite fauna associated with adult Stomoxys calcitmns in the U.K.

Abstract. Adult females of the blood-sucking muscid Stomoxys calcitrans sampled between June and September 1993 from a cattle farm ( n = 839) and from a pig farm ( n = 542) in North-West England were examined for mites. Twelve species of mites from ten families and three orders were identified as follows. In the Prostigmata, Eryenetes sp., Family Ereynetidae and Pediculaster mesembrinae , Family Pygmephoridae. In the Astigmata, Procalvolia zacheri Family Saproglyphidae, Acarusfarris , Family Acaridae, Bonomoia sphaerocerae and Myianoetus sp., Family Anoetidae. In the Mesostigmata, Macrocheles muscaedomesticae and Macrocheles subbadius Family Macrochelidae, Digamasellus sp., Family Digamasellidae, Halolaelaps sp., Family Halolaelapidae. Prodinychus sp., Family Uropodoidea and Thinoseius sp., Family Eviphididae. Mean infestation rates at the two sites (all mite species) for the entire sampling period were 31.6 ± 13.9% and 19.8 ± 3.6% respectively. 51 % of synbovine flies sampled in July were infested with mites. Mean numbers of flies infested in August at both farms were significantly lower compared to other months. The presence of tritonymphs of Ereynetes sp. on S.calcitrans demonstrates for the first time that this life cycle stage is naturally associated with insects in the field.
All mites were recovered from the ventral thorax and abdomen, and two or more species commonly infested individual flies. Associations of mites with their dipteran hosts are described and discussed.     

Effects of carbon dioxide, acetone and 1-octen-3-ol on the activity of the stable fly, Stomoxys calcitmns

Abstract. The responses of Stomoxys calcitrans (L.) to carbon dioxide, acetone and 1-octen-3-ol were assessed using flight activity as a measure of activation. Carbon dioxide and acetone caused significant increases in activity, with thresholds at -0.006% and -0.01 μg 1^-l, respectively. For l-octen-3-ol, flight activity decreased at 2 μg 1^-1 for males, and at 0.2 μg 1^-1 for females. Variation in activity was also manifest as differences in the time elapsed between landing and subsequent take-off: CO₂ (7.1 s) and acetone (12.2s) had lower times than the corresponding no-odour controls (16.6 and 23.2s), whereas 1-octen-3-ol (25 s) had a higher time than the control (21.5 s). The proportion of the total number of flights landing on a black target was higher in CO₂(0.16) and acetone (0.11) than in clear air (c. 0.07), but was lower for l-octen-3-ol.     

Diethylphenylacetamide: a new insect repellent against stable fly,Stomoxys calcitrans

Abstract. This paper reports the results of a laboratory study showing effectiveness of a new insect repellent N,N-diethylphenylacetamide (DEPA) against stable fly, Stomoxys calcitrans, and compared to N,N-diethyl-m-toluamide (DEET) and dimethyl-phthalate (DMP). DEPA gave maximum protection time of more than 6h at 3% concentration followed by DEET and DMP.     

Feeding response of the stable fly, Stomoxys calcitrant L., to blood fractions and adenine nucleotides

Abstract Feeding responses of the stable fly ( Stomoxys calcitrans L., Diptera: Muscidae) to blood fractions and saline-based diets were studied in an artificial feeding device. Flies fed equally on whole blood, plasma and erythrocyte fractions while resuspended platelets or the buffy layer did not stimulate feeding. Evidence from filtration studies indicates that plasma contains both less-than-5000D and greater-than-100,000D phagostimulatory components. ATP, ADP and AMP in saline stimulate feeding in this insect in a dose-dependent manner. cAMP also stimulates feeding in the range of concentrations tested. It is argued that the less-than-5000D phagostimulant may be one or more of these adenine nucleotides.