Zuccagnia punctata Cav. Essential Oil into Poly(ε-caprolactone) Matrices as a Sustainable and Environmentally Friendly Strategy Biorepellent against Triatoma infestans (Klug) (Hemiptera, Reduviidae).

The main strategies against Triatoma infestans (primary vector responsible for the Chagas disease transmission) are the elimination or reduction of its abundance in homes through the application of insecticides or repellents with residual power, and environmental management through the improvement of housing. The use of plant-derived compounds as a source of therapeutic agents (i.e., essential oils from aromatic plants and their components) is a valuable alternative to conventional insecticides and repellents. Essential oil-based insect repellents are environmentally friendly and provide reliable personal protection against the bites of mosquitoes and other blood-sucking insects. This study investigates, for the first time to our knowledge, the potential repellent activity of Zuccagnia punctata essential oil (ZEO) and poly(ε-caprolactone) matrices loaded with ZEO (ZEOP) prepared by solvent casting. The analysis of its essential oil from aerial parts by GC-FID and GC-MS, MS allowed the identification of 25 constituents representing 99.5% of the composition. The main components of the oil were identified as (-)-5,6-dehydrocamphor (62.4%), alpha-pinene (9.1%), thuja-2, 4 (10)-diene (4.6%) and dihydroeugenol (4.5%). ZEOP matrices were homogeneous and opaque, with thickness of 800 ± 140 µm and encapsulation efficiency values above 98%. ZEO and ZEOP at the lowest dose (0.5% wt./wt., 96 h) showed a repellency of 33 and 73% respectively, while at the highest dose (1% wt./wt., 96 h) exhibited a repellent activity of 40 and 66 %, respectively. On the other hand, until 72 h, ZEO showed a strong repellent activity against T. infestans (88% repellency average; Class V) to both concentrations, compared with positive control N-N diethyl-3-methylbenzamide (DEET). The essential oils from the Andean flora have shown an excellent repellent activity, highlighting the repellent activity of Zuccagnia punctata. The effectiveness of ZEO was extended by its incorporation in polymeric systems and could have a potential home or peridomiciliary use, which might help prevent, or at least reduce, Chagas' disease transmission.

Zuccagnia punctata Cav. Essential Oil into Poly(ε-caprolactone) Matrices as a Sustainable and Environmentally Friendly Strategy Biorepellent against Triatoma infestans (Klug) (Hemiptera, Reduviidae)

The main strategies against Triatoma infestans (primary vector responsible for the Chagas disease transmission) are the elimination or reduction of its abundance in homes through the application of insecticides or repellents with residual power, and environmental management through the improvement of housing. The use of plant-derived compounds as a source of therapeutic agents (i.e., essential oils from aromatic plants and their components) is a valuable alternative to conventional insecticides and repellents. Essential oil-based insect repellents are environmentally friendly and provide reliable personal protection against the bites of mosquitoes and other blood-sucking insects. This study investigates, for the first time to our knowledge, the potential repellent activity of Zuccagnia punctata essential oil (ZEO) and poly(ε-caprolactone) matrices loaded with ZEO (ZEOP) prepared by solvent casting. The analysis of its essential oil from aerial parts by GC­FID and GC-MS, MS allowed the identification of 25 constituents representing 99.5% of the composition. The main components of the oil were identified as (−)-5,6-dehydrocamphor (62.4%), alpha-pinene (9.1%), thuja-2, 4 (10)-diene (4.6%) and dihydroeugenol (4.5%). ZEOP matrices were homogeneous and opaque, with thickness of 800 ± 140 µm and encapsulation efficiency values above 98%. ZEO and ZEOP at the lowest dose (0.5% wt./wt., 96 h) showed a repellency of 33 and 73% respectively, while at the highest dose (1% wt./wt., 96 h) exhibited a repellent activity of 40 and 66 %, respectively. On the other hand, until 72 h, ZEO showed a strong repellent activity against T. infestans (88% repellency average; Class V) to both concentrations, compared with positive control N-N diethyl-3-methylbenzamide (DEET). The essential oils from the Andean flora have shown an excellent repellent activity, highlighting the repellent activity of Zuccagnia punctata. The effectiveness of ZEO was extended by its incorporation in polymeric systems and could have a potential home or peridomiciliary use, which might help prevent, or at least reduce, Chagas' disease transmission.

Analysis of Chagas disease vectors occurrence data: the Argentinean triatomine species database.

BACKGROUND: Chagas disease is a neglected tropical disease and Trypanosoma cruzi (its etiological agent) is mainly transmitted by triatomines (Hemiptera: Reduviidae). All triatomine species are considered as potential vectors; thus, their geographic distribution and habitat information should be a fundamental guide for the surveillance and control of Chagas disease. Currently, of the 137 species distributed in the Americas (Justi and Galvão 2017), 17 species are cited for Argentina: Panstrongylus geniculatus, P. guentheri, P. megistus, P. rufotuberculatus, Psammolestes coreodes, Triatoma breyeri, T. delpontei, T. eratyrusiformis, T. garciabesi, T. guasayana, T. infestans, T. limai, T. patagonica, T. platensis, T. rubrofasciata, T. rubrovaria and T. sordida. Almost 20 years have passed since the publication of the "Atlas of the Triatominae" by Carcavallo et al. (1998) and no work has been done to provide an updated complete integration and analysis of the existing information for Argentinean triatomine species. Here we provide a detailed temporal, spatial and ecological analysis of updated occurrence data for triatomines present in Argentina. NEW INFORMATION: This is the first database of the 17 triatomine species present in Argentina (15917 records), with a critical analysis of the temporal, spatial and ecological characteristics of 9788 records. The information spans the last 100 years (1918-2019) and it was mostly obtained from the DataTri database and from the Argentinean Vector Reference Center. As 70% of the occurrences corresponded to the last 20 years, the information was split into two broad periods (pre-2000 and post-2000). Occurrence data for most species show distribution range contractions, which, from the pre-2000 to post-2000 period, became restricted mainly to the dry and humid Chaco ecoregions. Concurrently, the highest species richness foci occurred within those ecoregions. The species T. infestans, T. sordida, T. garciabesi and T. guasayana mostly colonise human dwelling habitats. This study provides the most comprehensive picture available for Argentinean triatomine species and we hope that any knowledge gaps will encourage others to keep this information updated to assist health policy-makers to make decisions based on the best evidence.

Activity levels of female Triatoma infestans change depending on physiological condition.

BACKGROUND: Triatoma infestans (Hemiptera: Reduviidae) is the main vector of Trypanosoma cruzi, the etiological agent of Chagas disease, in South America. Active dispersal of this vector is the principal cause of recolonization of human dwellings previously treated with insecticides. Due to the persistence of vector populations and their movement between habitats, dispersive behavior studies are important for understanding the epidemiology of Chagas disease. The aim of this study was to analyze the relationship among T. infestans females' activity levels according to their physiological conditions. RESULTS: Two groups of insects were used, unfed and fed females. Each was composed of three subgroups in relation to the reproductive state: fifth-stage nymphs, virgin and fertilized females. There was a significant interaction between reproductive and nutritional states among T. infestans female' activity levels. During the experiments, unfed and fed nymphs remained inactive. Virgin females showed a dual behavior in their movement; fasted insects were more active. Fertilized females, both fed and unfed, were always active. CONCLUSION: The reproductive and nutritional conditions of T. infestans females affect their activity levels. When females with different reproductive states remained together, fertilized females showed permanent activity levels, suggesting that this subgroup of females represents the highest epidemiological risk as colonizers of human dwellings.

The process of lipid storage in insect oocytes: The involvement of ß-chain of ATP synthase in lipophorin-mediated lipid transfer in the chagas' disease vector Panstrongylus megistus (Hemiptera: Reduviidae).

Lipophorin is the main lipoprotein in the hemolymph of insects. During vitellogenesis, lipophorin delivers its hydrophobic cargo to developing oocytes by its binding to non-endocytic receptors at the plasma membrane of the cells. In some species however, lipophorin may also be internalized to some extent, thus maximizing the storage of lipid resources in growing oocytes. The ectopic ß chain of ATP synthase (ß-ATPase) was recently described as a putative non-endocytic lipophorin receptor in the anterior midgut of the hematophagous insect Panstrongylus megistus. In the present work, females of this species at the vitellogenic stage of the reproductive cycle were employed to investigate the role of ß-ATPase in the transfer of lipids to the ovarian tissue. Subcellular fractionation and western blot revealed the presence of ß-ATPase in the microsomal membranes of the ovarian tissue, suggesting its localization in the plasma membrane. Immunofluorescence assays showed partial co-localization of ß-ATPase and lipophorin in the membrane of oocytes as well as in the basal domain of the follicular epithelial cells. Ligand blotting and co-immunoprecipitation approaches confirmed the interaction between lipophorin and ß-ATPase. In vivo experiments with an anti-ß-ATPase antibody injected to block such an interaction demonstrated that the antibody significantly impaired the transfer of fatty acids from lipophorin to the oocyte. However, the endocytic pathway of lipophorin was not affected. On the other hand, partial inhibition of ATP synthase activity did not modify the transfer of lipids from lipophorin to oocytes. When the assays were performed at 4°C to diminish endocytosis, the results showed that the antibody interfered with lipophorin binding to the oocyte plasma membrane as well as with the transfer of fatty acids from the lipoprotein to the oocyte. The findings strongly support that ß-ATPase plays a role as a docking lipophorin receptor at the ovary of P. megistus, similarly to its function in the midgut of such a vector. In addition, the role of ß-ATPase as a docking receptor seems to be independent of the enzymatic ATP synthase activity.

Control of pyrethroid-resistant Chagas disease vectors with entomopathogenic fungi.

BACKGROUND: Triatoma infestans-mediated transmission of Tripanosoma cruzi, the causative agent of Chagas disease, remains as a major health issue in southern South America. Key factors of T. infestans prevalence in specific areas of the geographic Gran Chaco region-which extends through northern Argentina, Bolivia, and Paraguay-are both recurrent reinfestations after insecticide spraying and emerging pyrethroid-resistance over the past ten years. Among alternative control tools, the pathogenicity of entomopathogenic fungi against triatomines is already known; furthermore, these fungi have the ability to fully degrade hydrocarbons from T. infestans cuticle and to utilize them as fuel and for incorporation into cellular components. METHODOLOGY AND FINDINGS: Here we provide evidence of resistance-related cuticle differences; capillary gas chromatography coupled to mass spectrometry analyses revealed that pyrethroid-resistant bugs have significantly larger amounts of surface hydrocarbons, peaking 56.2+/-6.4% higher than susceptible specimens. Also, a thicker cuticle was detected by scanning electron microscopy (32.1+/-5.9 microm and 17.8+/-5.4 microm for pyrethroid-resistant and pyrethroid-susceptible, respectively). In laboratory bioassays, we showed that the virulence of the entomopathogenic fungi Beauveria bassiana against T. infestans was significantly enhanced after fungal adaptation to grow on a medium containing insect-like hydrocarbons as the carbon source, regardless of bug susceptibility to pyrethroids. We designed an attraction-infection trap based on manipulating T. infestans behavior in order to facilitate close contact with B. bassiana. Field assays performed in rural village houses infested with pyrethroid-resistant insects showed 52.4% bug mortality. Using available mathematical models, we predicted that further fungal applications could eventually halt infection transmission. CONCLUSIONS: This low cost, low tech, ecologically friendly methodology could help in controlling the spread of pyrethroid-resistant bugs.

Strong host-feeding preferences of the vector Triatoma infestans modified by vector density: implications for the epidemiology of Chagas disease.

BACKGROUND: Understanding the factors that affect the host-feeding preferences of triatomine bugs is crucial for estimating transmission risks and predicting the effects of control tactics targeting domestic animals. We tested whether Triatoma infestans bugs prefer to feed on dogs vs. chickens and on dogs vs. cats and whether vector density modified host choices and other vital rates under natural conditions. METHODOLOGY: Two host choice experiments were conducted in small caged huts with two rooms between which bugs could move freely. Matched pairs of dog-chicken (six) and dog-cat (three) were assigned randomly to two levels of vector abundance and exposed to starved bugs during three nights. Bloodmeals from 1,160 bugs were tested by a direct enzyme-linked immunosorbent assay. PRINCIPAL FINDINGS: Conditional logistic regression showed that dogs were highly preferred over chickens or cats and that vector density modified host-feeding choices. The relative risk of a bug being blood-engorged increased significantly when it fed only on dog rather than chicken or cat. Bugs achieved higher post-exposure weight at higher vector densities and successive occasions, more so if they fed on a dog rather than on a cat. CONCLUSIONS: Our findings strongly refute the hypothesis that T. infestans prefers to blood-feed on chickens rather than dogs. An increase in dog or cat availability or accessibility will increase the rate of bug feeding on them and exert strong non-linear effects on R(0). When combined with between-dog heterogeneities in exposure, infection, and infectiousness, the strong bug preference for dogs can be exploited to target dogs in general, and even the specific individuals that account for most of the risk, with topical lotions or insecticide-impregnated collars to turn them into baited lethal traps or use them as transmission or infestation sentinels based on their immune response to Trypanosoma cruzi or bug salivary antigens.

Effects of topical application of fipronil spot-on on dogs against the Chagas disease vector Triatoma infestans.

We assessed the insecticidal effects of fipronil spot-on applied to experimental dogs on the blood-feeding success and other vital parameters of the Trypanosoma cruzi vector Triatoma infestans. In the first trial, the cumulative mortality of 30 third or fourth instar nymphs exposed to eight fipronil-treated dogs differed significantly from those exposed to untreated dogs at 1 week post-treatment, but not at baseline or at 2-6 weeks post-treatment. In the second trial, the effects of multiple exposures to fipronil-treated dogs on bug population dynamics were assessed. A population of 80-84 bugs of various life stages were allowed to colonize eight closed experimental huts, and then exposed twice weekly to control or treated dogs over a period of 110 days and censused at monthly intervals. Throughout the trial, multiple exposure to fipronil did not significantly affect bug population size, fecundity, hatching, molting, survival, blood-feeding success and degree of engorgement. Only when engorgement was taken to include only fully fed bugs, did fipronil significantly reduce their degree of engorgement relative to bugs exposed to control dogs. We conclude that at tested dosages fipronil spot-on would have little effect in controlling (peri)domestic Tri. infestans or protecting dogs from contact with the bugs.

Extinction of experimental Triatoma infestans populations following continuous exposure to dogs wearing deltamethrin-treated collars.

Dogs are domestic reservoir hosts of Trypanosoma cruzi, the etiological agent of Chagas disease. We evaluated the effect of deltamethrin-treated dog collars (DTDCs) over time on the population dynamics of Triatoma infestans, a main T. cruzi vector. Forty founder bugs of mixed life stages were allowed to colonize mud-thatched experimental huts and exposed continuously to either uncollared control dogs (N = 3) or dogs wearing DTDCs (N = 7) for a period of up to 196 days. When compared with bugs exposed to control dogs, bugs exposed to collared dogs were shown to have reduced feeding success (odds ratio [OR] = 0.40; 95% confidence interval [CI], 0.26-0.63; P < 0.001) and lower survival (OR = 0.15; 95% CI, 0.08-0.29; P < 0.001); in fact, all of the bug populations exposed to collared dogs became extinct 77-196 days after study initiation. Bugs exposed to DTDC-wearing dogs were also shown to have a lower fecundity (i.e., number of eggs produced per live female bug: OR = 0.64; 95% CI, 0.51-0.81; P < 0.001) and molting rate to first-instar nymphs (OR = 0.32; 95% CI, 0.13-0.75; P < 0.01) than those bugs exposed to control dogs. DTDCs could represent a novel tool to prevent and control canine and (hence) human Chagas disease.

Distribution and pathogenicity of Trypanosoma cruzi isolated from peridomestic populations of Triatoma infestans and Triatoma guasayana from rural Western Argentina.

We assessed the distribution of Trypanosoma cruzi infection in peridomestic triatomines collected manually at a district-wide scale in rural villages around Olta, Western Argentina, and typed the isolated strains according to their pathogenicity to laboratory mice. Of 1623 triatomines examined, only 14 (0.9%) were infected with T. cruzi based on microscopical examination of feces. The prevalence of T. cruzi infection was 0.8% in Triatoma infestans, 2.3% in T. guasayana, and nil in T. garciabesi, T. platensis, and T. eratyrusiformis. Local transmission occurred in kitchens, store-rooms and goat corrals or nearby, though at very low levels. T. cruzi was detected by at least one parasitological method in 11 (79%) of 14 microscope-positive bugs. Hemoculture was the most sensitive method (67%) followed by culture of organ homogenates, histopathology or xenodiagnosis of inoculated suckling mice (55-58%), and culture of microscope-positive bug feces (46%). The evidence suggests that most of the isolated T. cruzi strains would be myotropic type III. Our study establishes for the first time that peridomestic, microscope-positive T. guasayana nymphs were actually infected with T. cruzi, and may be implicated as a putative secondary vector of T. cruzi in domestic or peridomestic sites.

Chagas disease control: deltamethrin-treated collars reduce Triatoma infestans feeding success on dogs.

Dogs are domestic reservoir hosts of Trypanosoma cruzi, the aetiological agent of Chagas disease. Using an experimental set-up mimicking rural mud-and-thatch houses, we evaluated the effect of deltamethrin-treated dog collars on the feeding success and survival of Triatoma infestans, the main T. cruzi vector in Latin America. Seven collared and three uncollared control dogs were exposed to colonized T. infestans at day 0 (i.e. before attachment of collars), at 15 days, and then monthly for 3 months post collar attachment. Following overnight exposure to uncollared dogs, 96% (1473/1538) of bugs fed, of which 51% (746/1473) fully engorged. Feeding rates were significantly reduced on collared dogs for up to 1 month post collar attachment with the lowest rates of 91% (551/604) observed at day 30 (P<0.05). Amongst those bugs that fed, engorgement rates were significantly reduced on collared dogs throughout the trial, during which average rates were 31% (543/1768) (P<0.001). No collar effect on individual bug survival was observed. Although observed effects on feeding and engorgement were limited, the strong association between blood-feeding, blood meal size and T. cruzi transmission suggests that deltamethrin-treated dog collars could help to control canine (and possibly human) T. cruzi infection.

Distribution and pathogenicity of Trypanosoma cruzi isolated from peridomestic populations of Triatoma infestans and Triatoma guasayana from rural Western Argentina

We assessed the distribution of Trypanosoma cruzi infection in peridomestic triatomines collected manually at a district-wide scale in rural villages around Olta, Western Argentina, and typed the isolated strains according to their pathogenicity to laboratory mice. Of 1623 triatomines examined, only 14 (0.9 percent) were infected with T. cruzi based on microscopical examination of feces. The prevalence of T. cruzi infection was 0.8 percent in Triatoma infestans, 2.3 percent in T. guasayana, and nil in T. garciabesi, T. platensis, and T. eratyrusiformis. Local transmission occurred in kitchens, store-rooms and goat corrals or nearby, though at very low levels. T. cruzi was detected by at least one parasitological method in 11 (79 percent) of 14 microscope-positive bugs. Hemoculture was the most sensitive method (67 percent) followed by culture of organ homogenates, histopathology or xenodiagnosis of inoculated suckling mice (55-58 percent), and culture of microscope-positive bug feces (46 percent). The evidence suggests that most of the isolated T. cruzi strains would be myotropic type III. Our study establishes for the first time that peridomestic, microscope-positive T. guasayana nymphs were actually infected with T. cruzi, and may be implicated as a putative secondary vector of T. cruzi in domestic or peridomestic sites.

Effectiveness of residual spraying of peridomestic ecotopes with deltamethrin and permethrin on Triatoma infestans in rural western Argentina: a district-wide randomized trial.

OBJECTIVE: To compare the effectiveness of a single residual spraying of pyrethroids on the occurrence and abundance of Triatoma infestans in peridomestic ecotopes in rural La Rioja. METHODS: A total of 667 (32.8%) peridomestic sites positive for T. infestans in May 1999 were randomly assigned to treatment within each village, sprayed in December 1999, and reinspected in December 2000. Treatments included 2.5% suspension concentrate (SC) deltamethrin in water at 25 mg active ingredient (a.i.)/m(2) applied with: (a) manual compression sprayers (standard treatment) or (b) power sprayers; (c) 1.5% emulsifiable concentrate (EC) deltamethrin at 25 mg a.i./m(2); and (d) 10% EC cis-permethrin at 170 mg a.i./m(2). EC pyrethroids were diluted in soybean oil and applied with power sprayers. All habitations were sprayed with the standard treatment. FINDINGS: The prevalence of T. infestans 1-year post-spraying was significantly lower in sites treated with SC deltamethrin applied with manual (24%) or power sprayers (31%) than in sites treated with EC deltamethrin (40%) or EC permethrin (53%). The relative odds of infestation and catch of T. infestans 1-year post-spraying significantly increased with the use of EC pyrethroids, the abundance of bugs per site before spraying, total surface, and host numbers. All insecticides had poor residual effects on wooden posts. CONCLUSION: Most of the infestations probably originated from triatomines that survived exposure to insecticides at each site. Despite the standard treatment proving to be the most effective, the current tactics and procedures fail to eliminate peridomestic populations of T. infestans in semiarid rural areas and need to be revised.

Flight metabolism in Panstrongylus megistus (Hemiptera: Reduviidae): the role of carbohydrates and lipids

The metabolism of lipids and carbohydrates related to flight activity in Panstrongylus megistus was investigated. Insects were subjected to different times of flight under laboratory conditions and changes in total lipids, lipophorin density and carbohydrates were followed in the hemolymph. Lipids and glycogen were also assayed in fat body and flight muscle. In resting insects, hemolymph lipids averaged 3.4 mg/ml and significantly increased after 45 min of flight (8.8 mg/ml, P < 0.001). High-density lipophorin was the sole lipoprotein observed in resting animals. A second fraction with lower density corresponding to low-density lipophorin appeared in insects subjected to flight. Particles from both fractions showed significant differences in diacylglycerol content and size. In resting insects, carbohydrate levels averaged 0.52 mg/ml. They sharply declined more than twofold after 15 min of flight, being undetectable in hemolymph of insects flown for 45 min. Lipid and glycogen from fat body and flight muscle decreased significantly after 45 min of flight. Taken together, the results indicate that P. megistus uses carbohydrates during the initiation of the flight after which, switching fuel for flight from carbohydrates to lipids.

Flight metabolism in Panstrongylus megistus (Hemiptera: Reduviidae): the role of carbohydrates and lipids.

The metabolism of lipids and carbohydrates related to flight activity in Panstrongylus megistus was investigated. Insects were subjected to different times of flight under laboratory conditions and changes in total lipids, lipophorin density and carbohydrates were followed in the hemolymph. Lipids and glycogen were also assayed in fat body and flight muscle. In resting insects, hemolymph lipids averaged 3.4 mg/ml and significantly increased after 45 min of flight (8.8 mg/ml, P < 0.001). High-density lipophorin was the sole lipoprotein observed in resting animals. A second fraction with lower density corresponding to low-density lipophorin appeared in insects subjected to flight. Particles from both fractions showed significant differences in diacylglycerol content and size. In resting insects, carbohydrate levels averaged 0.52 mg/ml. They sharply declined more than twofold after 15 min of flight, being undetectable in hemolymph of insects flown for 45 min. Lipid and glycogen from fat body and flight muscle decreased significantly after 45 min of flight. Taken together, the results indicate that P. megistus uses carbohydrates during the initiation of the flight after which, switching fuel for flight from carbohydrates to lipids.