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Insects sense thermal cues mainly through thermoreceptors located in the antenna. To analyse the impact of antennectomy on the thermal behaviour of the haematophagous bug Triatoma infestans, we studied the distribution of intact and antennectomised bugs in an experimental arena where a temperature gradient was established, as well as the biting response of insects with and without antennae to objects at the temperature of a potential host. Antennectomy did not abolish thermopreference, but modified the temperature at which the insects preferred to stay. In the arena, antennectomised insects chose to remain at a higher temperature (ca. 3 °C higher in unfed bugs), and exhibited a larger dispersion around that preferred temperature, than intact bugs. In addition, ablated insects temporarily lost their ability to bite an object at the temperature of a potential host, but that ability was gradually recovered after the fifth day post-antennectomy. Results presented here show that thermoreceptors other than those located on the antennae can also guide thermal behaviours. We conclude that the function of antennal thermoreceptors can be taken over by other receptors located in different regions of the body. Those receptors have a different sensitivity and confer the insects with a different responsiveness.

International audience; The adaptation to light of compound eyes in insects has been extensively documented and their adaptive role is well understood. Much less attention has been paid, however, to the control of ocelli sensitivity, a study which could help us to understand the functional role of these simple eyes. We analyzed the dynamic changes in the distribution of screening pigments which occur in the ocelli of the haematophagous bug, Triatoma infestans, when the insects are subjected either to light/dark cycles (LD), to constant darkness (DD) or constant light (LL). We then compared these changes with those occurring in the compound eyes of the same individuals and found that, while compound eyes are subject to the control of an endogenous circadian clock, the adaptation of the ocelli is entirely dependent on environmental illumination. In addition, we have observed that environmental temperature is not involved in the control of screening pigments in either ocelli or compound eyes as a direct stimulus, nor as a Zeitgeber. The existence of a differential control in the components of the dual visual system represents an adaptive advantage in the adjustment of visual sensitivity in insects exposed to quick changes in lighting conditions in their natural habitat. We discuss the implications of our findings with regards to the biology of triatomines and with respect to the general understanding the functional role of insect ocelli.

Compounds from the metasternal and Brindley's glands of the blood-sucking bug, Triatoma infestans, were identified by solid phase microextraction (SPME) and gas chromatography-mass spectrometry. Volatile compounds released by adult bugs during copulation or after mechanical disturbance were also characterized. Six compounds were identified and found consistently in all samples from metasternal glands. The most abundant were 3-pentanone, 2-methylbutanol, 3-pentanol, and an unidentified compound. The metasternal gland blends did not differ qualitatively between sexes. Compounds found in Brindley's glands were short chain acids, alcohols, esters, and a ketone with no qualitative differences between sexes. Isobutyric acid was the main component of this blend, and two new confirmed compounds were described as products of these glands: 2-butanone and 2-methylbutyric acid. 3-Pentanone was collected from the headspace over 33% of the copulating pairs of T. infestans. Volatiles found in the headspace of disturbed T. infestans adults included short-chain fatty acids, alcohols, esters, and ketones, with no qualitative differences between sexes. Both types of glands apparently discharge their contents after disturbance. However, most of the volatiles released by bugs after disturbance came from Brindley's glands. The locomotor activity of fourth instars increased significantly after stimulation with the odors emitted by disturbed adults, as compared with larvae stimulated by the odor of undisturbed adults or by clean air. We also studied the directional behavioral response of fifth instars to the disturbance scent in a locomotion compensator. Larvae exposed to volatiles released by disturbed adults walked away from the direction of the odor. The results suggest that this blend or part of it functions as an alarm pheromone for T. infestans. We suggest that the metasternal glands of this species are involved both in the sexual and the alarm contexts, and that the Brindley's glands probably have both alarm and defensive roles.

We studied the spectral sensitivity of the visual system of the blood-sucking bug Triatoma infestans, one of the main vectors of Chagas Disease in South America. We quantified the photonegative reaction of this insect in a rectangular arena, half of which was kept dark and the other half illuminated with various intensities of different monochromatic lights (or broadband stimuli for lambda665 nm). As a behavioral parameter of the photonegative response, we measured the time each insect spent in the dark half of the arena. We found that low intensity levels (under 0.06 microW/cm(2)) of monochromatic lights of 397, 458, 499, and 555 nm evoked a statistically significant (i.e., different from that of control groups) photonegative reaction. Insects were less sensitive to monochromatic lights of 357 nm (UV) and 621 nm (dark orange), and to broadband stimuli in the red part of the spectrum (665-695 nm). These findings indicate that the visual system of T. infestans is sensitive to broader regions of the spectrum than those previously reported.

The role of short-chain fatty acids in the host-seeking behaviour of Triatoma infestans larvae was investigated using a locomotion compensator. Several short-chain fatty acids were tested alone over a wide range of doses, or in combination with L-lactic acid (L-LA; 100 microg). Bugs showed no attractive response to single carboxylic acids, but when L-LA was added to airstreams carrying specific intensities of either propionic (C3; 100 microg), butyric (C4; 1 microg) or valeric acid (C5; 1 microg), these mixtures elicited an attractive response, evincing a synergistic effect. No orientation response was observed when caproic acid (C6) was offered with L-LA at the doses tested. Two blends were created: (1) C3, C4 and C5 combined at the effective doses when added with L-LA [C3C4C5 (1)], and (2) C3, C4 and C5 combined at a third of those intensities [C3C4C5 (2)]. Both blends were tested alone, with L-LA (100 microg), with a sub-threshold concentration of CO(2) (300 p.p.m. above the ambient level), and combined with both compounds together. Oriented responses of bugs were only observed with the blend (2) added with L-LA and with the combination of this lure with CO(2). This last combination evoked a behavioural response similar in intensity to that induced by a live mouse.

Behavioural responses of Triatoma infestans larvae to carbon dioxide and other odours of vertebrate origin were investigated in a locomotion compensator. T. infestans oriented towards airstreams enriched with carbon dioxide exhibiting a threshold response between 300 and 400 p.p.m. above the ambient CO(2) background. The accuracy of the oriented response to carbon dioxide improved with stimulus intensity. Remarkably, insects did not show any change in their sensitivity threshold to carbon dioxide with the starvation time. The attractiveness to carbon dioxide depended on the time of the day, i.e. these nocturnal bugs only oriented towards carbon dioxide-loaded airstreams during the first hours of the scotophase. L-lactic acid did not evoke oriented responses when it was presented as a single stimulus in a wide range of intensities. However, a marked synergism was evident when L-lactic acid was combined with a sub-threshold concentration of carbon dioxide. Under this condition, the threshold response to carbon dioxide decreased to 75-150 p.p.m. above ambient CO(2) background. The isomer D-lactic acid evoked no response, either alone or in combination with carbon dioxide. When insects were stimulated with 1-octen-3-ol a significant positive orientation was found. This response was not modified by the addition of carbon dioxide.

The temporal modulation of the behavioural response to carbon dioxide and its chronobiological basis were investigated in larvae of Triatoma infestans. We analysed the orientation towards CO(2) of insects kept under three different illumination regimes: (1) 12 h light/12 h darkness cycles (L/D), (2) constant darkness (D/D) and (3) constant light (L/L). When maintained under L/D conditions, insects exhibited an oriented response towards airstreams added with 1500 ppm of CO(2) during the first hours of the scotophase only. Bugs maintained under D/D also showed a positive orientation response towards CO(2) during the first hours of the subjective night, while bugs kept under L/L did not show a rhythmic oriented behaviour. Thus, T. infestans displayed a daily rhythm of orientation towards CO(2) (i.e. a potential food source) only at the beginning of the scotophase. The persistence of the rhythm under constant darkness reveals the existence of an endogenous circadian control of this behaviour.

The chronobiological basis of the daily dynamics of thermopreference was tested in adults of Triatoma infestans, continuously registering the preferred temperature of insects released over a temperature gradient for 13 days. We found that the thermopreference in T. infestans is a dynamic process that depends on the time of the day and the post-feeding time. When submitted to a 12:12 h light/darkness cycle (L/D), the preferred temperature reached the highest and the lowest values at the end of the light and dark phases, respectively. This daily rhythm persisted under constant conditions of illumination (D/D and L/L), suggesting the existence of an internal oscillator controlling this behaviour. Statistical analysis revealed that the thermopreference of insects kept under L/D exhibited a ca. 24 h periodicity, while insects kept in D/D and L/L showed free-running periods of τ(D/D)=23.35 and τ(L/L)=27.35 h, respectively. The persistence of a cyclic pattern of thermopreference under constant conditions, and free-running periods, close to, but different from 24 h, demonstrate the existence of an endogenous control of the thermopreference in this species. The biological relevance of these results is discussed in the light of the hypothesis that both the length of time elapsed since feeding and the time of the day modify thermopreference in these bugs. The gradual decline in preferred temperature following feeding may be associated with energy conservation during starvation. The rhythmic modulation of thermopreference may be associated with the daily rhythm of locomotion activity shown by these bugs.

The behavioural response to water vapour of the haematophagous bug Triatoma infestans was analysed. Dry or humid discrete sources at different temperatures were used as stimuli for insects walking on a locomotion compensator. Humidity significantly increased the tendency of these bugs to orientate towards thermal sources. Furthermore, humid sources at room temperature were attractive to T. infestans, but this effect was limited to short-range distances. On the other hand, dynamic sources, i.e. airstreams carrying different water vapour contents did not affect the spontaneous anemotactic behaviour of this species, neither in sign (positive) nor in intensity. The anemotactic behaviour was also not influenced by the physiological water balance state of the bugs. Results are discussed in relation to the cues released by living hosts of triatomine bugs and in relation to their responses to air-currents.

SUMMARY We analysed dynamic changes in the ommatidial structure of the compound eyes of Triatoma infestans. This nocturnal insect possesses open-rhabdom eyes, in which a ring of six rhabdomeres from retinula cells 1–6 (R1–6) surrounds a central pair of rhabdomeres from retinula cells 7 and 8 (R7–8). Screening pigments are located in all the photoreceptors and in the primary (PPC) and secondary (SPC) pigment cells. During the day, pigments within R1–6 and the PPCs form a small ‘pupil’ above the rhabdom and pigments within R7–8 are clustered around the central rhabdomere, allowing light to reach only the central rhabdomere. At night, the ‘pupil’ widens, and pigments inside R7–8 concentrate in the proximal region of the cells, allowing light to reach the peripheral rhabdomeres. In addition, the distance between the cornea and the rhabdom decreases. These rhythmic changes adapt the sensitivity of the eye by controlling the amount of light reaching and travelling within the rhabdom. Furthermore, the rhythm persists under conditions of constant darkness (DD), i.e. it is controlled by an endogenous oscillator. Remarkably, there are differences in pigment movements between the retinula cells of a single ommatidium. The migration of pigments in R1–6 is regulated by a circadian input, while that in R7–8 is regulated by both direct light and circadian inputs. The rhythm vanishes under constant-light conditions (LL). In this species, the circadian rhythm of photonegative behaviour persists in both DD and LL conditions, suggesting that these two rhythms, in retinal morphology and visual behaviour, may be generated by different circadian oscillators.

We analysed how the assembling behaviour of Triatoma infestans is modulated by the convergence of chemical cues released by their faeces and the spectral quality of the light associated with refuges. Second-instar larvae were confronted with refuges associated with a visual stimulus (either blue, green or red lights having the same intensity, or darkness) and a chemical cue (presence or absence of faeces). In this context, faeces constitute a major attractant for bugs. In the absence of faeces, bugs always assembled in dark places. Green light was always rejected despite the presence of faeces, i.e. the assembling behaviour was controlled by a photonegative reaction to this light. In the presence of red light, orientation towards the chemical cue dominated over the bugs' photonegative reaction to this light. Such a light was avoided in the absence of faeces but not in their presence. Also, negative phototaxis to blue light could be counteracted by the presence of the chemical cue. Thus, a concrete interaction between visual and olfactory cues occurred in these experiments: the bugs' response changed depending on the specific combination of spectral light and faeces. Finally, bugs responded differentially to blue, red and green lights of the same intensity. They assembled preferentially on red, followed by blue and then by green. Thus, discrimination between lights of different spectral quality is possible, probably through an achromatic mechanism.

The simple eyes (ocelli) of recently emerged adult Triatoma infestans exhibit a narrow elongated "pupil", surrounded by a ring of brown-reddish pigment, the "iris". This pupil does not respond to changes in the illumination, but varies in size after the imaginal ecdysis. This change corresponds, internally, with the growth of the corneal lens and the associated retina up to an age of about 20 days. This has not been previously observed in an insect. The use of this characteristic for recognising young adults of this species is suggested.

The effectiveness of a trap for triatomines baited with yeast cultures has been previously demonstrated for Triatoma infestans in laboratory assays. We report here results from laboratory assays testing yeast traps for Triatoma sordida, Triatoma brasiliensis, Triatoma pseudomaculata, and Panstrongylus megistus. All assays were conducted in an open experimental arena 100 cm X 100 cm, with two traps placed at opposite sides of the arena. One of the traps contained a yeast culture, and the other trap contained a saccharose solution as a control. Two series of experiments were done, one without a refuge for the insects and one with a refuge. The results obtained clearly demonstrated that the yeast-baited traps were effective in the laboratory in capturing both T. sordida and P. megistus. For T. sordida, yeast-baited traps captured significantly more bugs than did the control traps (t test P value = 0.03). For P. megistus, when a refuge was provided during the assay, yeast-baited traps also captured significantly more bugs than did the control traps (t test P value = 0.006). In the experiments with T. brasiliensis and T. pseudomaculata, both traps captured some insects, but the yeast traps captured many fewer bugs than was true with the T. sordida and P. megistus bugs. These results indicate that, in the laboratory, yeast traps can capture considerable numbers of T. sordida and P. megistus in one night. We discuss the potential use of yeast traps for detecting and capturing both triatomine species.

The projections of mechanosensory hairs located on the dorsal and lateral head of the adult haematophagous bug Triatoma infestans were analyzed by means of cobalt filling. Axons run into the anterior and posterior tegumentary nerve and project through the brain to the ventral nerve cord. The fibres are small in diameter and run as a fascicle. Some branches run into suboesophageal and prothoracic centres; others run as far as to the mesothoracic ganglion. These sensory projections resemble that of wind-sensitive head hairs of the locust. The functional role of this sensory system in this species is discussed.

The negative phototactic response of Triatoma infestans (Hemiptera) was studied in an arena, half of which was kept dark and the other half illuminated with different light intensitites. For each intensity we measured the time the insects spent in the dark half, the time to reach the side opposite to that where they were released, and the number of passages through the middle line of the arena. T. infestans displayed a photonegative behaviour that was enhanced by high light intensities. Bugs maintained in 12:12 light-dark cycles responded differently to the same illumination levels when tested in their photophase and scotophase: sensitivity to light was higher during the latter. Bugs entrained to light-dark cycles, kept afterwards either in constant darkness or in constant light, and tested in their subjective night and day, showed the same responses as bugs from the light-dark group tested in their corresponding photophase and scotophase. Thus, phototactic sensitivity is under endogenous control. The behaviour shown by T. infestans may be understood as being composed of at least two different drives: an exploratory one, and a negative phototactic response that is under endogenous control and is particularly sensitive to light during the scotophase, when activity peaks occur.

The response of Triatoma infestans (Klug) to odours left behind by walking insects was analysed. The insects aggregated on papers marked with putative chemical footprints in the absence of other assembling chemical signals. Contact of the receptor insect with the treated surface was necessary to elicit the response because the signal was not able to attract insects, only to arrest them on marked surfaces. These requirements contrast with other chemical aggregation signals of triatomines that attract the insect and involve olfactory responses. By washing the cuticle of the insects with hexane and impregnating papers with the extract, a response was evoked in test insects similar to that elicited by papers on which insects had walked. The biological role of this novel aggregation factor is discussed in relation to other assembling signals of different origin.

Triatominae bugs aggregate around their faeces which carry an assembly signal. Here the biological role of the signal is studied for Triatoma infestans . We describe a Stereotypic behaviour of T. infestans where the insects exit from shelters in order to defaecate. A higher concentration of faecal depositions are found at accesses to refuges. In choice experiments where faeces were presented in association with artificial refuges to ascertain whether this affected election of shelters by the bugs, shelters with faeces, either inside or outside, were significantly preferred by bugs. We conclude that faeces help in the selection of refuges by these insects and the role of excrement as a chemical signal for a shelter under use is discussed.

The projections of first-order ocellar interneurons were analyzed in the hematophagous bug Triatoma infestans by cobalt filling. The axons run between the calyces of the mushroom bodies and dorsal of the central body to different regions of the brain and the subesophageal and thoracic ganglia. The interneurons can be grouped into large L cells and small S cells. The L cells have cell bodies ranging from 11.5 to 25 μm and axons ranging from 8 to 25 μm diameter (measured in the ocellar nerve); the S cells have smaller cell bodies of 9 μm or less and axon diameters less than 5 μm. The projections of ten L cells are described in detail; they project to the protocerebral posterior slope (PS), the other ocellus (O), the optic neuropile, and the subesophageal, pro-, meso-, and metathoracic ganglia, either to ipsi- (PS I, II), or contra- (PS IV, V), or bilateral areas. In this case projections occur to the same areas (PSO, PS III) or different areas at each side (PSOE; E = eye). Large-descending (LD) first-order interneurons project to the contralateral posterior slope of the protocerebrum, the deutocerebrum, and subesophageal, pro-, mesa-, and metathoracic areas (LD I-III). Cell bodies are located in the dorsal protocerebral lobes and pars intercerebralis, except the PS II neuron and three LD cells, which are located in the ipsilateral posterior protocerebrum. This is the first report about ocellar pathways in Hemiptera. Their adaptive function is discussed with reference to the bugs' behavior as Chagas disease vectors. © 1996 Wiley-Liss, Inc.

The behaviour of Triatoma infestans (Hemiptera: Reduviidae) in response to thermal stimulation was studied. We analysed the effect of a heat source on the locomotion activity, and antennal movements of tethered insects. Antennal movements were studied both in the horizontal and vertical planes, and related to whether locomotion was being displayed or not. In addition, we analysed the trajectories of intact and unilaterally antennectomised insects towards a warm cylinder in a closed-loop design. In the presence of a thermal source, triatomine bugs performed more antennal movements and walked faster than in its absence. Insects displayed a sequence of walking bouts and standing phases of variable length, concomitant with active antennal movements in both the horizontal and vertical planes. While walking, the antennae moved smoothly in co-ordination. When the insects stopped, the movements changed dramatically, displaying abrupt changes of direction. The trajectories of control animals were straight towards the thermal source. Unilateral antennectomy resulted in deviated trajectories towards the goal, with the deviation always occurring towards the side of the intact antenna. The trajectories were fairly straight up to the moment of the extension of the proboscis, at a fixed distance of around 15 mm between the tip of the head and the warm surface, when a sharp body turn occurred. Control animals probed the source at a position perpendicular to the warm surface. Antennectomised animals missed the goal and probed the air at a position tangential to the surface of the warm body. The function of the antennae during the approach to a potential source of food, and the mechanisms of orientation possibly involved, are discussed.

The effect of the addition of egg yolk to a blood meal on the development of Triatoma infestans (Klug) was analysed. Egg yolk was added in different quantities to heparinized, citrated or defibrinated bovine blood, which was offered in an artificial feeder. Control groups were fed on either non-supplemented blood or hen in vivo. Results indicated that the addition of egg yolk to a blood meal reduced the intermoult interval and the moult success increased when egg yolk was added to the diet. The effect was more dramatic for the less suitable diets, as follows: citrated blood > defibrinated blood > heparinized blood. The minimal and most effective concentration of egg yolk in the diet was 10% v/v. When the egg yolk was centrifuged in a density-gradient, fractions with different performance could be obtained; middleweight fractions, but not the heaviest or lightest ones reproduced the observed effects of the whole yolk at the corresponding concentration. Growth rate and moulting success showed to be differentially affected by the diets and the growth performance of in vivo-fed experimental groups could be completely reproduced by adding yolk to the blood diets offered in artificial feeders.

Assembling behaviour associated with mating was investigated in Triatoma infestans. The spatial distribution of both sexes was observed by video films, in the presence or absence of a copulating pair. Males aggregated around copulating pairs. Females did not exhibit this behaviour and their mean spatial density remained unaffected. Spontaneous aggregation tendency was observed in males in the absence of a copulating pair, but the temporal course significantly differed from that observed in the presence of a mating pair. Results support the existence of an aggregation signal that is released during mating, affecting the behaviour of males.

In the second segment of the antennae of haematophagous reduviids an unusual cave-like organ is found the function os which was investigated in Triatoma infestans. the morphology of the organ makes it difficult to ascribe it to a mechno- or chemoreceptive function, but shows some characteristics shared with thermoreceptors of other animals. The electrical activity of sense cells was recorded in the presence of stimuli that evoke behavioural responses in this species, i.e. warm, CO2, lactic and butyric acids at different concentrations. The three compounds tested failed to evoke a response at all concentrations assayed. Only thermal stimulation evinced a clear modification in the electrical activity of the sense cells.Both the morphological and electrophysiological findings support a thermoreceptive finding, habitat selection and circadian synchronization.

The hygropreference of larvae and adults of the reduviid bug Triatoma infestans (Klug) was investigated along relative humidity (RH) gradients established by means of different salt solutions, as well as by H 2 SO 4 dilutions. Both larvae and adults preferred to remain at or near 0% RH. This preference was similar for bugs reared in the laboratory under different conditions, and for those collected in the field. Several assays to control for both experimental asymmetries along the gradient and the effects of thigmotactic behaviour permit exclusion of these factors as responsible for the behaviour of the insects. The preferred RH did not vary with the time of the day or fasting. Larval insects exhibited a significant tendency to remain close to 0% RH at the time of ecdysis. The same preference was exhibited to ovipositing females. The success of ecdysis and egg hatching were measured at different RH values ranging from 0 to 75%. No differences were found between groups; eclosion success ranged from 86 to 94% for egg-hatching and from 89.5 to 95% for ecdysis, at all RHs tested. Results agree with high saturation deficits found along the geographical areas exhibiting the highest prevalence of T. infestans . However, they are in contradiction with values usually recommended to rear this species in the laboratory. The adaptive value of the hygropreference is discussed.

Triatominae bugs remain quiet during the day, when they can be usually found assembled in protected sites, becoming active at night. By means of olfactometers and in different experimental arenas, we studied in the laboratory the response of Triatoma infestans (Klug) to volatiles present in their faeces and the dynamics of the aggregation behaviour. In a choice arena, nymphs showed a significant tendency to aggregate around papers impregnated with dry faeces. In a simultaneous discrimination olfactometer, and air current passed by a paper impregnated with dry faeces also elicited a significant attraction. Freshly deposited faeces did not elicit attraction, but rather a significant rejection was observed. Immediately after feeding, insects did not prefer impregnated papers and were randomly distributed in the arena. The change in the response from indifference to attraction takes place within the first 24 h after feeding. Finally, a spontaneous daily rhythm of aggregation/dispersion was demonstrated from the 6th day after feeding onwards. Maxima of assembling and dispersion occur at the onset of the photophase and scotophase, respectively.

The response to intra- and interspecific faecal assembling signals was tested in Rhodnius prolixus. Papers impregnated with excrement of R. prolixus induced the aggregation of larvae of this species, but also of those of Triatoma infestans. However, faeces belonging to T. infestans were not able to assemble larvae of R. prolixus. On the other hand, there was no response of R. prolixus to putative chemical factors from their cuticle (footprints), in contrast to T. infestans. Results are discussed as related to the ecology of both species.

Daily locomotor activity of Triatoma infestans adults exhibits two major components, one at the second hour of the scotophase and a second one at the beginning of the photophase. The occurrence of peaks can be ascribed to food and refuge search at dusk and dawn, respectively. Both maxima appear under discrete light-dark regimes (12 h light-12 h dark, 25:0 lx) at constant temperature, as well as under discrete ligh-low temperature cycles 12 h at 30°C-12 h at 20°C in continuous light or in darkness. Under constant darkness, both maxima of activity display different freerunning periods (τa ≈ 23 h 50 min, τb ≈ 22 h 15 min), suggesting the involvement of two oscillators controlling individually, the two activity peaks. Under continuous light, however, activity becomes unimodal, freerunning at τ ≈ 26 h 40 min. Resetting a 180° shift of high-low temperature cycles occurs faster than by light-dark cycles. In conditions of zeitgeber competition, by simultaneous exposition to light and temperature cycles at various phase relationships, light preponderantly controls the synchronization of activity. Results indicate that the activity of T. infestans is controlled by a circadian system involving a hierarchical arrangement of coupled oscillators. Environmental temperature modulates the activity level and the time course of the spontaneous locomotion. Antennectomy affects the temporal expression of the activity, insects becoming active at an unusual daytime.

The thermal preference of adults of the reduviid bug Triatoma infestans (Klug) was investigated along a temperature gradient by means of video recording. The preferred temperature varied with a daily rhythm, reaching the highest value at the onset and the lowest at the end of the dark phase. The daily range of preferred temperatures depended on the starvation level. After feeding it was about 28–29°C, and after 12 days of starvation 25–26°C. The number of eggs laid showed a bimodal pattern, with maxima at 29°C and 26°C. Along a gradient of relative humidity, the bugs were always found around 40% r.h.

The gorging response of 3rd-instar nymphs of the vector of Chagas' disease Triatoma infestans Klug to different diets was analysed using an artificial feeder which allows one to feed a great number of bugs simultaneously. Citrated blood was more acceptable than heparinated blood, either stored at 4°C or at –20°C. Freezing of blood at –20°C and their subsequent haemolysis did not affect the intake of citrated blood when it was defrosted immediately before the experiment, but in fact when this occurred 36 hs before. The addition of mice odours to the artificial feeder did not improve the reaction of the bugs to the food it offer. Saline solutions without phagostimulants proved to evoke gorging. Zusammenfassung Die Zucht von Triatoma infestans King (Het., Reduviidae) ohne lebendes Wirtstier. I. Einflusse auf die aufgenommene Futtermenge Es wurde die Blutaufnahme von Nymphen im 3. Stadium von Triatoma infestans, dem Ubertrager der Chagas-Krankheit, untersucht. Geboten wurden verschiedene Diaten in einer kunstlichen Futterquelle, die die gleichzeitige Futterung einer grosen Anzahl von Wanzen ermoglichte. Bei 4°C als auch bei –20 °C konserviertes Citratblut wurde besser akzeptiert als Heparinblut. Die Konservierung des Blutes bei –20°C (d. h. Hamolysis) hat die Akzeptanz des Blutes dann nicht beeinflust, wenn es kurz vor der Futterung aufgetaut wurde. Lag der Auftauzeitpunkt langer zuruck (36 h) wurde von den Tieren weniger Blut aufgenommen. Der Zusatz von Duft von Mausen an der Futterquelle verbesserte die Aufnahme nicht. Die Tiere nahmen auch reine Salzlosungen ohne Phagostimulantien auf.

Carbon dioxide is generally recognized as an important cue used by haematophagous insects to locate a food source. When the mammalian hosts of these insects breathe, they normally emanate considerable amounts of CO2 at discrete intervals, i.e. with each exhalation. In this work, we analysed the effect of temporally pulsing CO2 on the host-seeking behaviour of Triatoma infestans. We investigated the ability of T. infestans to follow continuous and intermittent air pulses of 0.25, 0.5 and 1 Hz that included different concentrations of CO2. We found that insects were attracted to pulsed airstreams of 0.25 and 0.5 Hz transporting 400 ppm of CO2 above the ambient levels and to continuous streams added with the same amount of CO2. On the other hand, insects walked away from streams pulsed at rates of 1 Hz regardless of the amount of CO2 they bear. The walking trajectories displayed by bugs to attractive CO2-pulsed streams were as rectilinear and accurate as those to CO2-continuous streams. Our results are discussed in the frame of the interaction between olfactory and mechanoreceptive inputs as affecting the behavioural response of bugs.

We analysed the flying activity of Triatoma infestans and Rhodnius prolixus when confronted to artificial lights of different spectral quality. We found that the presence of light sources (white or ultraviolet) did not affect their spontaneous take-off rate. The comparison between species showed that R. prolixus was more prone to fly than T. infestans. Females of T. infestans initiated flight more frequently than males of the same species. Although the same tendency was observed in R. prolixus, no significant differences were assessed between sexes. Concerning the orienting behaviour of triatomines at take-off in relation to the position of the light source, T. infestans showed a significant tendency to fly towards white light, but a non-oriented response when confronted to UV light or in the absence of a light source. R. prolixus also preferred to fly towards a source of white light and exhibited a non-oriented response with no light. However, when the UV light was presented, these bugs exhibited a bimodal attraction/repellence-behaviour. Our results support true attraction by white light rather than menotaxis or arrival by chance. These findings are discussed in relation to the colonization of human dwellings by Chagas disease vectors.

Triatominae bugs experience changes in the mechanical properties of their cuticle prior to feeding. This process-plasticization-allows a rapid stretching of the unsclerotized abdominal cuticle of triatominae larvae and it is evoked by sensory inputs related to feeding. We tested: (a) whether the cuticle recovers its original mechanical properties after plasticization, (b) whether repeated stimulation would be able to evoke recurrent plasticization along the same larval instar, (c) the temporal course of recovering cuticular stiffness. We injected Ringer solution into the body cavity of the bugs at constant pressure, using the injection rate (ml/min) as a measure of the cuticle extensibility. To trigger plasticization, individuals were allowed to feed on blood from an artificial feeder at 32+/-2 degrees C. After plasticization occurred, the abdominal cuticle gradually recovered its original mechanical properties. Bugs were capable of plasticizing for a second time when repeatedly stimulated. The effects of plasticization vanished between 1 and 2 h after stimulation. Although one full meal could suffice to accomplish moult in other Triatomine species, Triatoma infestans is able to feed repeatedly during a single larval instar. Accordingly to this, their cuticle recovers stiffness in some hours and becomes able to respond repeatedly to sensory inputs associated with feeding.

We tested the aggregation response to inter and intraspecific chemical signals in Panstrongylus megistus and Triatoma infestans. As previously described for T. infestans, larvae of P. megistus significantly aggregated on papers impregnated with their own excrement and on papers marked with cuticular substances deposited on surfaces on which these insects had walked. T. infestans bugs also aggregated on papers impregnated by faeces or by cuticular substances of P. megistus, and P. megistus aggregated on papers contaminated by faeces or by cuticular substances of T. infestans. The response of P. megistus to its cuticular substances was significantly stronger than that to its faeces. The non-specificity of the two signals is discussed in the context of the ecological relationship between both species.

Background Triatomine bugs are the insect vectors of Trypanosoma cruzi, the etiological agent of Chagas disease. These insects are known to aggregate inside shelters during daylight hours and it has been demonstrated that within shelters, the aggregation is induced by volatiles emitted from bug feces. These signals promote inter-species aggregation among most species studied, but the chemical composition is unknown. Methodology/Principal Findings In the present work, feces from larvae of the three species were obtained and volatile compounds were identified by solid phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS). We identified five compounds, all present in feces of all of the three species: Triatoma infestans, Panstrongylus megistus and Triatoma brasiliensis. These substances were tested for attractivity and ability to recruit insects into shelters. Behaviorally active doses of the five substances were obtained for all three triatomine species. The bugs were significantly attracted to shelters baited with blends of 160 ng or 1.6 µg of each substance. Conclusions/Significance Common compounds were found in the feces of vectors of Chagas disease that actively recruited insects into shelters, which suggests that this blend of compounds could be used for the development of baits for early detection of reinfestation with triatomine bugs., Author Summary Chagas disease is a parasitic infection affecting approximately 12 million people, and is considered to be one of the most severe burdens for public health in Latin America. Control of the disease is based on attempted elimination of domestic populations of triatomine bugs, the insects transmitting the disease to humans, by means of insecticide spraying. Currently, vigilance programs monitoring triatomine reinfestation processes in houses are performed by manual search for bugs. Effective and sustainable new methods allowing continuous monitoring of domestic triatomine populations are required. Based on the fact that the insects hide in dark refuges that are marked by volatile signals emitted in their feces, we screened the feces of three species for volatile compounds common to these prominent vectors. The potential for these odors to promote triatomine aggregation was evaluated and we present evidence that a synthetic blend of these substances is capable of recruiting bugs into shelters, mimicking the natural pheromone. This blend may be used to develop a bait to monitor triatomine reinfestation processes in a similar manner as is used commonly for the monitoring of agricultural pests.

Simple eyes or ocelli coexist with compound eyes in many adult insects. The change in the morphology of the ocelli along the five larval instars of Triatoma infestans was studied by light and scanning electron microscopy. Our analysis showed that the development of the ocelli of these bugs occurs gradually along the larval life. The photoreceptor layer is present from the second-instar onwards. The cornea appears first at the imaginal stage and grows up to the 18-20th day after the last ecdysis, associated to an increase in the retinal mass. Findings are discussed in a comparative fashion and in relation to the functionality of the ocellar system in T. infestans.

Adults and larvae of Triatoma infestans spend daylight hours assembled in shaded places. An assembling factor has been demonstrated in the excrement of this species. We analysed different aspects of the dynamics of the response of bugs. Recently fed insects do not aggregate around faeces. They start to show a significant assembling response from the 8th hour after feeding onwards. Just deposited faeces do not evoke assembling, but a significant rejection instead. This reaction switches 3 h after deposition, when the faeces become attractive to the insects. The attractiveness of faeces persists for about 10 days and can be recovered after this time by rehydration. These findings are discussed in relation to the biological role of faeces and the dynamics of the use of refuges by T. infestans.

The preference of Triatoma infestans for shelters at different temperatures or relative humidities (RH) was tested in laboratory assays. The insects preferred refuges at a temperature of 34 degrees C rather than 26 degrees C, and chose to stay in refuges with an RH of 20% rather than 80%. Temperature and RH records made inside and outside experimental chicken-houses under natural climatic conditions showed that such habitats tend to maintain an inner climate favourable to T. infestans. Typical refuges in adobe walls showed strongly damped fluctuations in minimum and maximum temperatures, and delayed changes in these parameters when compared with an external reference. Similarly, the jarilla (Larrea sp.), a plant used for constructing roofing, damps fluctuations in minimum and maximum RH and delays these RH fluctuations when compared to an external reference. We discuss possible effects of environmental factors on the distribution of insects in human dwellings, and analyse the environmental characteristics of normally available refuges in materials used for house building in endemic zones.

The pattern of locomotion activity was studied in Triatoma infestans, in relation to the use of an artificial refuge offered to the bugs in an experimental arena. In assays performed with insects that had a post-ecdysis starvation period of 1 week, the activity was low and mainly related to refuge leaving or entering. Insects that had gone through a longer period of starvation (6 weeks) exhibited a high locomotory activity throughout the night. Specific temporal windows were detected for refuge leaving and entering after dusk and before dawn, which were not modified by different levels of starvation of the insects. T. infestans exhibits a higher motivation for feeding during the first night hours, in comparison to that observed at the end of the scotophase. The activity peaks previously reported for T. infestans are therefore related to host and refuge search.

Factors affecting mating behaviour in the bug Triatoma infestans were quantitatively studied in the laboratory. Experimental conditions were established so that the probability of copulation increased with the time elapsed since the first adult meal. Copulatory attempts by males did not vary as a function of time, but rejections by females became significantly less frequent with the post feeding time. Non-receptive females displayed four types of rejection behaviour, i.e. flattening, stridulation, evasion and abdominal movements. The occurrence of stridulation performed by females in a sexual context was observed in a regular fashion and was quantified for the first time in this species.

The timing of hatching of Triatoma infestans (Klug) eggs was analyzed by means of time-lapse photography. Under a photoperiod of 12:12 (L:D) and constant darkness, maximum eclosion occurred at the end of the subjective night. Under constant light, the rhythmicity disappeared. Hatching rhythm seems to be maintained by a circadian endogenous oscillator. By hatching close to dawn, when relative humidity is high, insects may avoid excessive water loss.

Triatoma bugs turn away from a vertical black stripe on the inner surface of a rotating or oscillating drum by keeping it in the lateral visual field at an angle α relative to the long axis of the body. The value of α depends on the angular velocity w of the drum. Below w=5° s−1, α increases with increasing w, and the stripe can lie to either side of the animal. Occasionally, the bugs switch between these two tracking modes. Above w=5° s−1, α remains nearly constant at about 120° and the stripe lags behind the animal. We call this lateral tracking. At velocities over 5° s−1 the animals track the leading edge of a wide black stripe in the same manner as they track a narrow stripe. Below 5° s−1 they walk towards the centre of the stripe (skototaxis). Objects moving towards the insect above the horizon are also fixated at an angle of about 120°. Lateral tracking is mediated mainly by the dorsal part of the visual field, as can be shown by occluding either the dorsal or ventral halves of the eyes. The walking speed of the bugs increases significantly during lateral tracking of an oscillating stripe compared with that during frontal fixation of a stationary one. We therefore interpret lateral tracking as an escape response.

The response of blood-sucking bugs ( Triatoma infestans ) to thermal stimulation, in the absence of warm-air gradients or convection currents, was analysed, using windows with different infrared transmittance. When some infrared radiation could reach the insects, they responded as without window. When infrared-opaque windows were used, no response was evoked. Discrimination of the source temperatures was tested using two sources at different temperatures and open-loop conditions. The bugs orientated toward that source that had the temperature of a food source and preferred it to a source at 50°C, in spite of both sources being 10 cm distant. From behavioural evidence we conclude that T. infestans perceives the radiant heat of a warm source and can estimate its temperature at a distance, when seeking for food.

Le dispositif experimental choisi permet de faire varier la temperature de surface du nourissoir de celle du sang qu'il contient. Le reflexe du rostre et la piqure dependent de la temperature de surface. La temperature du sang intervient sur le nombre d'insectes qui se nourissent.

It has been observed that Triatoma infestans and Rhodnius prolixus females stridulate to reject copulatory attempts performed by males. In addition, triatomines stridulate when disturbed or handled. In the present study, the temporal structure and frequency spectra of vibrational signals produced by mechanically disturbed T. infestans, T. sordida, T. guasayana, R. prolixus and Dipetalogaster maxima were analysed and compared. The inter-ridge distances of the prosternal stridulatory organ of the same species were also measured. The frequency spectra and repetition rates were similar, despite individuals of these five species have different sizes, their stridulatory grooves have different inter-ridge distances, and also their vibratory signals exhibited different temporal patterns. The hypothesis that disturbance stridulations are non-specific signals and could function to deter predators was discussed.

The effectiveness of a trap for triatomines baited with yeast cultures has been previously demonstrated in laboratory assays. We report results from assays testing yeast-traps for Triatoma infestans performed under natural climatic conditions. All assays were conducted at experimental chicken-coops colonised by bugs, situated at an endemic area for Chagas disease in Argentina. Two different models of traps were tested, for use either on the floor, or attached to walls. The results obtained clearly demonstrate that yeast-baited traps are effective for capturing triatomine bugs. Floor traps, when baited, captured significantly more bugs than their controls (t-test, P = 0.0008, k = 10). In addition, wall traps were also more effective, when compared to their controls (t-test, P = 0.011, k = 10). The maximum capture by a single yeast-trap was 39 bugs in one night, while for the same period the maximum capture by a control-trap was two bugs. Traps captured significantly more bugs at nights when maximum temperature was above 18 degrees C (t-test, P = 0.0002). Results indicate that yeast traps are capable of capturing significant numbers of free insects in one night. The potential use of yeast- traps for the detection of triatomine bugs is discussed.

In addition to compound eyes, most adult insects posses two or three simple eyes, the ocelli. The function of these photoreceptors remains elusive in most cases. Triatomine bugs posses two well-developed ocelli, located in a latero-dorsal position, behind the compound eyes. We tested the role of the ocelli in the phototactic behaviour of Triatoma infestans, by measuring the time spent by adult males in the dark half of an experimental arena, which had the other half illuminated. The occlusion of the ocelli or the compound eyes alone had little effect on the phototactic response of the bugs. Only those insects which had both their ocelli and compound eyes occluded showed a significant reduction in their negative response to light. The ability of the ocelli of T. infestans to mediate the phototactic response by themselves (i.e., not through the modulation of compound eyes sensitivity) constitutes the first report on this function in insects.