Spermathecae: Morphofunctional features and correlation with fat bodies and trachea in six species of vectors of Chagas disease.

Since spermatheca is able to transport spermatozoa and maintain a specific microenvironment for the storage of viable sperm cells for long periods of time, specific morphofunctional features must be involved in this capacity, and an efficient nutritional and oxygen supply must be required. In this study, we investigated the histological features of spermathecae and fat bodies in six species of three genera of epidemiological importance for Chagas' disease. The association of the reproductive system with the fat bodies and tracheal system was also focused in these species. The reproductive system, tracheae and fat bodies were fixed in 4% formaldehyde, and embedded in glycol methacrylate. The sections were stained with H.E., picrosirius red and Periodic-Acid Schiff methods for morphological analyses. Paraffin-embedded spermatheca sections were submitted to immunofluorescence for detection of V-ATPase. In P. lignarius, R. montenegrensis and R. prolixus, the spermatheca contains a slightly dilated tubular distal portion. In P. megistus and T. tibiamaculata, the spermatheca shows a large bulbous distal portion, and in T. infestans, a large oval-shaped distal portion. In all species, this portion was surrounded by a thin muscular layer, and the epithelial height varied according to the shape of this terminal portion. All spermathecal proximal portions showed simple columnar epithelium surrounded by a thick muscular layer. The epithelial cells of spermathecae showed PAS-positive cytoplasm and V-ATPase immunofluorescence in the apical surface. Tracheoles and polysaccharide-rich fat body cells were found next or in close contact to the oviduct or spermathecal tissues. The results indicate that the spermatheca proximal portion is related to contraction and sperm transport, whose oxygen and energy supply is guaranteed by the associated tracheal branches and fat bodies. In the storage portion, fat bodies and tracheae seem to be crucial for the maintenance of an optimal spermathecal microenvironment and storage of viable sperm cells. The participation of V-ATPase in the spermathecae epithelial cells may contribute for the maintenance of an optimal luminal milieu to spermatozoa, by alkalinization and/or acidification of lumen, similarly to the other epithelial cell types in insects. Further studies are necessary to clarify the role of this proton pump in the spermathecal epithelial cells.

Biological characteristics of Meccus phyllosomus pallidipennis (Hemiptera: Reduviidae: Triatominae) fed on two different hosts

Abstract INTRODUCTION: Different blood meal sources can affect biology of triatomines. METHODS: We studied the influence of two different blood meal sources (rabbits and hens) on five biological parameters in Meccus phyllosomus pallidipennis and collected biological parameters. RESULTS: The cohort fed on rabbits had a shorter median life-cycle and lower mortality rate than that fed on hens. Both cohorts required a similar number of blood meals to reach adulthood. Median longevity and number of blood meals for adults were similar between both cohorts. CONCLUSIONS The studied parameters reflect the high grade of adaptation of M. p. pallidipennis feeding on different hosts.

Parámetros biológicos de Meccus phyllosomus phyllosomus (Burmeister) 1835, de Triatoma recurva (Stål) 1868 (Hemiptera, Reduviidae) y de sus híbridos de laboratorio / Biological parameters of Meccus phyllosomus phyllosomus (Burmeister), 1835, Triatoma recurva (Stål), 1868 (Hemiptera, Reduviidae) and their laboratory hybrids

Resumen Introducción. Los híbridos de especies y subespecies de triatominos mexicanos exhiben características biológicas y de comportamiento sobresalientes comparadas con las de sus parentales, específicamente su vigor híbrido, lo cual puede incrementar el riesgo de transmisión de Trypanosoma cruzi a sus huéspedes humanos y animales. Objetivo. Describir y comparar los valores de los parámetros biológicos relacionados con el ciclo de vida de Triatoma recurva, de Meccus phyllosomus phyllosomus y de sus híbridos de laboratorio, para estimar su riesgo como transmisores de T. cruzi a las poblaciones humanas. Materiales y métodos. Se entrecruzaron ejemplares de T. recurva y de M. p. phyllosomus y se obtuvo una cohorte híbrida. Las tres cohortes se mantuvieron bajo condiciones similares de laboratorio. En las tres se compararon los datos de sus ciclos de vida, el número de alimentaciones necesarias para mudar, el porcentaje de mortalidad, el de eclosión de huevos y la fecundidad de las hembras. Resultados. La cohorte híbrida tuvo el menor tiempo promedio entre el primer instar o estadio y el adulto (136,6 ± 17,5 días). El número de alimentaciones para cambiar de instar (13,4 ± 1,7; 15,6 ± 5,2) fue mayor en la cohorte híbrida y en la de T. recurva. La cohorte híbrida registró la menor mortalidad (29 %) y el mayor porcentaje (70 %) de hembras al final del ciclo biológico. La fecundidad por hembra por día (rango: 1,1 ± 0,6 -1,6 ± 0,7) fue similar entre las poblaciones. El porcentaje de eclosión de huevos fue superior a 70 % en la cohorte híbrida y en la de M. p. phyllosomus. Conclusión. La cohorte híbrida presentó algunas características biológicas que implicarían un mayor poder transmisor de T. cruzi que el de sus grupos parentales.

Abstract Introduction: Hybrids from species and subspecies of Mexican triatomines have shown outstanding biological and behavioral characteristics as compared with their parental groups. This could lead to an increasing risk of transmission of Trypanosoma cruzi to human and animal reservoir hosts. Objective: To describe and compare biological parameters related to the life cycle of Triatoma recurva, Meccus phyllosomus phyllosomus and their laboratory hybrids and estimate their risk as transmitters of T. cruzi to humans. Materials and methods: We kept a cohort of each one of the three groups (T. recurva, M. p. phyllosomus and their hybrids) under similar laboratory conditions and then compared them to record the data related to their life cycles, number of blood meals to molt, mortality rates, egg eclosion rates and female fecundity. Results: The hybrid cohort showed the lowest mean time from first-instar nymph to adult (136.6 ± 17.5 days). The number of blood meals from molt to next instar (13.4 ± 1.7; 15.6 ± 5.2) was higher in the hybrid and T. recurva cohorts. The hybrid cohort showed the lowest mortality rate (29.5%) and the highest rate (70%) of adult females at the end of the biological cycle. Fecundity per day per female (range: 1.1 ± 0.6- 1.6 ± 0.7 eggs) was similar in the three cohorts. Egg hatching rate was higher than 70% in both hybrid and M. p. phyllosomus cohorts. Conclusion: The hybrid cohort showed some outstanding biological characteristics, which would entail a higher risk of transmission of T. cruzi to hosts than their parentals.

Description of the feeding preferences of triatominae in the Chagas disease surveillance study for the State of Pernambuco, Brazil (Hemiptera: Reduviidae)

Abstract INTRODUCTION: Studying the feeding preferences of triatomines is an important entomological surveillance tool, since continuous surveillance of the disease is necessary. METHODS: The precipitin reaction was used to describe the feeding preferences of triatomines along with their natural infection by flagellates similar to Tyrpanosoma cruzi. Six hundred eighty-seven insects were examined, including Triatoma brasiliensis, Triatoma pseudomaculata, and Panstrongylus lutzi. RESULTS: Sixty-nine (10%) of 687 triatomines examined tested positive for flagellates similar to T. cruzi, and 8 (1.2%) of these fed on human blood. CONCLUSIONS: This study found potential transmitters of Chagas disease both inside and outside the domiciliar environment.

On triatomines, cockroaches and haemolymphagy under laboratory conditions: new discoveries.

For a long time, haematophagy was considered an obligate condition for triatomines (Hemiptera: Reduviidae) to complete their life cycle. Today, the ability to use haemolymphagy is suggested to represent an important survival strategy for some species, especially those in genus Belminus. As Eratyrus mucronatus and Triatoma boliviana are found with cockroaches in the Blaberinae subfamily in Bolivia, their developmental cycle from egg to adult under a "cockroach diet" was studied. The results suggested that having only cockroach haemolymph as a food source compromised development cycle completion in both species. Compared to a "mouse diet", the cockroach diet increased: (i) the mortality at each nymphal instar; (ii) the number of feedings needed to molt; (iii) the volume of the maximum food intake; and (iv) the time needed to molt. In conclusion, haemolymph could effectively support survival in the field in both species. Nevertheless, under laboratory conditions, the use of haemolymphagy as a survival strategy in the first developmental stages of these species was not supported, as their mortality was very high. Finally, when Triatoma infestans, Rhodnius stali and Panstrongylus rufotuberculatus species were reared on a cockroach diet under similar conditions, all died rather than feeding on cockroaches. These results are discussed in the context of the ecology of each species.

On triatomines, cockroaches and haemolymphagy under laboratory conditions: new discoveries

For a long time, haematophagy was considered an obligate condition for triatomines (Hemiptera: Reduviidae) to complete their life cycle. Today, the ability to use haemolymphagy is suggested to represent an important survival strategy for some species, especially those in genus Belminus. As Eratyrus mucronatus and Triatoma boliviana are found with cockroaches in the Blaberinae subfamily in Bolivia, their developmental cycle from egg to adult under a “cockroach diet” was studied. The results suggested that having only cockroach haemolymph as a food source compromised development cycle completion in both species. Compared to a “mouse diet”, the cockroach diet increased: (i) the mortality at each nymphal instar; (ii) the number of feedings needed to molt; (iii) the volume of the maximum food intake; and (iv) the time needed to molt. In conclusion, haemolymph could effectively support survival in the field in both species. Nevertheless, under laboratory conditions, the use of haemolymphagy as a survival strategy in the first developmental stages of these species was not supported, as their mortality was very high. Finally, when Triatoma infestans, Rhodnius stali and Panstrongylus rufotuberculatus species were reared on a cockroach diet under similar conditions, all died rather than feeding on cockroaches. These results are discussed in the context of the ecology of each species.

Intrusive versus domiciliated triatomines and the challenge of adapting vector control practices against Chagas disease

Chagas disease prevention remains mostly based on triatomine vector control to reduce or eliminate house infestation with these bugs. The level of adaptation of triatomines to human housing is a key part of vector competence and needs to be precisely evaluated to allow for the design of effective vector control strategies. In this review, we examine how the domiciliation/intrusion level of different triatomine species/populations has been defined and measured and discuss how these concepts may be improved for a better understanding of their ecology and evolution, as well as for the design of more effective control strategies against a large variety of triatomine species. We suggest that a major limitation of current criteria for classifying triatomines into sylvatic, intrusive, domiciliary and domestic species is that these are essentially qualitative and do not rely on quantitative variables measuring population sustainability and fitness in their different habitats. However, such assessments may be derived from further analysis and modelling of field data. Such approaches can shed new light on the domiciliation process of triatomines and may represent a key tool for decision-making and the design of vector control interventions.

Ecoepidemiology and biology of Eratyrus mucronatus Stål, 1859 (Hemiptera: Reduviidae: Triatominae), a sylvatic vector of Chagas disease in the Brazilian Amazon

Introduction Eratyrus mucronatus Stål, 1859 is a wild triatomine vector of Trypanosoma cruzi Chagas, 1909. However, little is known regarding the biology and ecoepidemiology of this triatomine in the Brazilian Amazon. The present study describes the biology of E. mucronatus grown under laboratory conditions and the epidemiological aspects of its natural breeding sites. Methods Five colonies were monitored in the field for 3 years. Temperature and humidity measurements were taken in the mornings and afternoons at the natural breeding sites, and the behavior and distribution of the nymphs and adults were observed in the wild colony. We also monitored the life cycle under controlled laboratory conditions. Results Some factors that were considered decisive for the establishment of these colonies were present at all of the colonies studied in the field. These factors included an active termite nest, a vertebrate for repast, and dry and shaded substrates with temperatures of 24-28°C and with humidity of 80-90%. A generation was developed in 274 days under these microclimatic conditions in the laboratory. Conclusions The climatic variables described in the field indicate that these environmental parameters have a limiting effect on the dispersal and colonization of E. mucronatus to new environments. ...

Demographic fitness of Belminus ferroae (Hemiptera: Triatominae) on three different hosts under laboratory conditions

Triatominae are widely recognised for their role as vectors of Trypanosoma cruzi. One of the main biological characteristics of this subfamily is their obligate haematophagous condition. However, previous studies on Belminus herreri and Belminus ferroae suggested that cockroaches are their principal hosts in domiciles. Due to this peculiar behaviour, the aim of this study was to analyse several demographic and reproductive parameters of B. ferroae fed on three different hosts (mice, cockroaches and Rhodnius prolixus) and relate B. ferroae fitness to these alternative hosts. The cohorts were reared under constant conditions. The egg hatching rate was similar for cohorts fed on cockroaches (69.4%) and R. prolixus (63.8%), but was much lower for the cohort fed on mice (16%). The development time from the nymph to adult stage and the average age of first reproduction (α) presented lower values in the cohort fed on cockroaches, which is consistent with the higher population growth rate associated with this host. Demographic parameters [intrinsic rate of natural increase, finite rate of population growth, net reproductive rate and damping ratio] showed statistically significant differences between the cohorts. Analysis of the life history of B. ferroae revealed a higher fitness related to the cockroach. The implications of these results for the origin of the subfamily are discussed.

Demographic fitness of Belminus ferroae (Hemiptera: Triatominae) on three different hosts under laboratory conditions.

Triatominae are widely recognised for their role as vectors of Trypanosoma cruzi. One of the main biological characteristics of this subfamily is their obligate haematophagous condition. However, previous studies on Belminus herreri and Belminus ferroae suggested that cockroaches are their principal hosts in domiciles. Due to this peculiar behaviour, the aim of this study was to analyse several demographic and reproductive parameters of B. ferroae fed on three different hosts (mice, cockroaches and Rhodnius prolixus) and relate B. ferroae fitness to these alternative hosts. The cohorts were reared under constant conditions. The egg hatching rate was similar for cohorts fed on cockroaches (69.4%) and R. prolixus (63.8%), but was much lower for the cohort fed on mice (16%). The development time from the nymph to adult stage and the average age of first reproduction (α) presented lower values in the cohort fed on cockroaches, which is consistent with the higher population growth rate associated with this host. Demographic parameters [intrinsic rate of natural increase, finite rate of population growth, net reproductive rate and damping ratio] showed statistically significant differences between the cohorts. Analysis of the life history of B. ferroae revealed a higher fitness related to the cockroach. The implications of these results for the origin of the subfamily are discussed.

Less invasive blood sampling in the animal laboratory: clinical chemistry and haematology of blood obtained by the Triatominae bug Dipetalogaster maximus.

Dipetalogaster maximus (Dipmax), a blood-sucking bug belonging to the family Reduviidae, has been used to obtain blood samples, for example for clinical chemistry and haematology, in a variety of zoo animals and wildlife. Using this bug allows stress-free blood sampling as the bug is able to draw blood without the mammal noticing the bug. In laboratory animal science, the need for blood samples from unstressed animals may arise, especially in animal behaviour research. The use of Dipmax bugs may prove a valuable tool for this purpose. To validate the method, we compared an array of standard blood parameters sampled from New Zealand White rabbits, sampled either by the use of bugs or by the conventional method; puncture of vena auricularis caudalis. The overall hypothesis was that there was no significant difference in clinical chemistry and haematological parameters between the bug method and the conventional method. A total of 17 clinical parameters as well as 12 haematological parameters were measured and compared in New Zealand White rabbits. The results showed that for 13 of these 29 analysed parameters, the bug method and the conventional method did not give significantly different results, and the obtained results were thus directly comparable. For the remaining parameters the obtained results were significantly different. However, all parameters were measurable in the bug samples. The influences of the bug metabolism on these parameters are discussed.

Ecological patterns of blood-feeding by kissing-bugs (Hemiptera: Reduviidae: Triatominae)

Host use by vectors is important in understanding the transmission of zoonotic diseases, which can affect humans, wildlife and domestic animals. Here, a synthesis of host exploitation patterns by kissing-bugs, vectors of Chagas disease, is presented. For this synthesis, an extensive literature review restricted to feeding sources analysed by precipitin tests was conducted. Modern tools from community ecology and multivariate statistics were used to determine patterns of segregation in host use. Rather than innate preferences for host species, host use by kissing-bugs is influenced by the habitats they colonise. One of the major limitations of studies on kissing-bug foraging has been the exclusive focus on the dominant vector species. We propose that expanding foraging studies to consider the community of vectors will substantially increase the understanding of Chagas disease transmission ecology. Our results indicate that host accessibility is a major factor that shapes the blood-foraging patterns of kissing-bugs. Therefore, from an applied perspective, measures that are directed at disrupting the contact between humans and kissing-bugs, such as housing improvement, are among the most desirable strategies for Chagas disease control.

Biological and genetic aspects of crosses between species of the genus Meccus (Hemiptera: Reduviidae Triatominae)

The degree of reproductive isolation between Meccus phyllosomus and the remaining five species of the genus Meccus, as well as between Meccus bassolsae and Meccus pallidipennis, Meccus longipennis and Meccus picturatus, was examined. Fertility and the segregation of morphological characteristics were examined in two generations of hybrids from crosses between these species. The percentage of couples with offspring (fertile) was high in the vast majority of sets of crosses, with the exception of that between ♀M. phyllosomus and ♂Meccus mazzottii. In sets of crosses involving M. bassolsae specimens, no first-generation (F1) individuals were morphologically similar to M. bassolsae, but instead shared the morphology of the other parental species. A similar phenomenon was observed in most sets of crosses involving M. phyllosomus. These results indicated that different degrees of reproductive isolation exist among the species of Meccus involved in this study. The biological evidence obtained in this study does not support the proposal that M. bassolsae is a full species. It could indicate that, on the contrary, it should be considered a subspecies of a single polytypic species. The biological evidence does support the proposal that M. phyllosomus is a full species.

Attraction of Chagas disease vectors (Triatominae) to artificial light sources in the canopy of primary Amazon rainforest

Adult triatomines occasionally fly into artificially lit premises in Amazonia. This can result in Trypanosoma cruzi transmission to humans either by direct contact or via foodstuff contamination, but the frequency of such behaviour has not been quantified. To address this issue, a light-trap was set 45 m above ground in primary rainforest near Manaus, state of Amazonas, Brazil and operated monthly for three consecutive nights over the course of one year (432 trap-hours). The most commonly caught reduviids were triatomines, including 38 Panstrongylus geniculatus, nine Panstrongylus lignarius, three Panstrongylus rufotuberculatus, five Rhodnius robustus, two Rhodnius pictipes, one Rhodnius amazonicus and 17 Eratyrus mucronatus. Males were collected more frequently than females. The only month without any catches was May. Attraction of most of the known local T. cruzi vectors to artificial light sources is common and year-round in the Amazon rainforest, implying that they may often invade premises built near forest edges and thus become involved in disease transmission. Consequently, effective Chagas disease prevention in Amazonia will require integrating entomological surveillance with the currently used epidemiological surveillance.

Absence of domestic triatomine colonies in an area of the coastal region of Ecuador where Chagas disease is endemic

Rhodnius ecuadoriensis is considered the second most important vector of Chagas disease in Ecuador. It is distributed across six of the 24 provinces and occupies intradomiciliary, peridomiciliary and sylvatic habitats. This study was conducted in six communities within the coastal province of Guayas. Triatomine searches were conducted in domestic and peridomestic habitats and bird nests using manual searches, live-bait traps and sensor boxes. Synantrhopic mammals were captured in the domestic and peridomestic habitats. Household searches (n = 429) and randomly placed sensor boxes (n = 360) produced no live triatomine adults or nymphs. In contrast, eight nymphs were found in two out of six searched Campylorhynchus fasciatus (Troglodytidae) nests. Finally, Trypanosoma cruzi DNA was amplified from the blood of 10 percent of the 115 examined mammals. Environmental changes in land use (intensive rice farming), mosquito control interventions and lack of intradomestic adaptation are suggested among the possible reasons for the lack of domestic triatomine colonies.

Searching for triatomines. A new method for field search using UV light.

Detection of triatomine bugs within a house is essential for the estimation of Chagas disease transmission risk and for evaluating the success of insecticidal control attempts. Small residual populations could represent an important risk but are difficult to detect by time manual sampling. Faecal marks from triatomines are clearly detectable with an ultraviolet (UV) light on most of the materials frequently used in rural buildings. A new method for finding triatomines is proposed here, based on the unexplored property of faeces to fluoresce when exposed to UV light.

Feeding behaviour of Belminus ferroae (Hemiptera: Reduviidae), a predaceous Triatominae colonizing rural houses in Norte de Santander, Colombia.

Belminus ferroae (Hemiptera: Reduviidae) was collected from houses in the localities of San Alberto and Santa Catalina in the municipality of Toledo, Department of Norte de Santander, Colombia. Precipitin tests revealed that the main hosts of B. ferroae are insects belonging to the order Blattodea, whose haemolymph serves as food (89.43%). A low rate of haematophagy was indicated on rodents, dogs and humans (7.32%) and no natural infection with Trypanosoma cruzi was found by either microscopic examination or polymerase chain reaction. The finding of B. ferroae inside houses, feeding primarily on Blattodea with only a minor tendency to feed on vertebrate blood, is indicative of both the evolutionary path from predator to blood-sucker taken by Triatominae and their tendency to adapt from silvatic to domestic habitats.

Occurrence of hybrids and laboratory evidence of fertility among three species of the Phyllosoma complex (Hemiptera: Reduviidae) in Mexico

In seven studied communities of Western Mexico, triatomine specimens were sympatrically collected, some with atypical morphological characteristics in contrast to pure specimens, which were presumed to be hybrids. More than 200 specimens of Meccus pallidipennis and Meccus longipennis with brown-yellow markings on dorsal connexival segments were collected in Ahuacapán and Quitupan. In La Mesa, more than 60 specimens similar to Meccus picturatus in most morphological characteristics (including size) were collected, although they presented a largely yellowish corium like M. pallidipennis. Interfertility was proven between all of the studied wild hybrid specimens, as well as between all the experimental laboratory hybrids. Two different phenotypes (M. picturatus and M. longipennis) were obtained from crosses between M. picturatus x M. picturatus and M. longipennis x M. longipennis from the three studied localities in state of Nayarit as from La Mesita. Results support the hypothesis that the subspecific ranking of those triatomines may, therefore, be more appropriate because reproductive isolation has not been developed and complete interbreeding was recorded.

Exploiting triatomine behaviour: alternative perspectives for their control

Living in close association with a vertebrate host and feeding on its blood requires different types of adaptations, including behavioural adjustements. Triatomines exhibit particular traits associated with the exploitation of their habitat and food sources and these traits have been the subject of intense analysis. Many aspects of triatomine behaviour have been relatively well characterised and some attempts to exploit the behaviours have been undertaken. Baited traps based on host-associated cues, artificial refuges and light-traps are some of the tools used. Here we discuss how our knowledge of the biology of Chagas disease vectors may help us sample and detect these insects and even increase the efficiency of control measures.

Sylvatic triatominae: a new challenge in vector control transmission

Over the last 10 years, Uruguay, Chile and Brazil have been certified as being free from disease transmission by Triatoma infestans, the main domiciliated vector for Chagas disease in the Southern Cone countries. This demonstrates that programmes addressing the vector for the disease's transmission are effective. These programmes have resulted in a dramatic decrease in the incidence of Chagas disease in Latin America. Guatemala was certified a few months ago as being free from disease transmission by Rhodnius prolixus, the main domiciliated vector for Chagas disease in Central American countries. However, the main concern for different countries' current control programmes is the continuity and sustainability of future vector control actions. The prevalence and incidence figures for individuals infected by Trypanosoma cruzi in Mexico and Andean and Central American countries highlights the need for broadened strategies in the struggle against the disease and its vectors. A number of triatomine insects are parasite vectors, each with a different life history. Therefore, it is important that new vector control strategies be proposed, keeping in mind that some species are found in peridomiciliary areas and wild ecotopes. The only viable control strategy is to reduce human interactions with vector insects so that the re-infestation and re-colonisation of human habitats will not take place.