New structures for goat corrals to control peridomestic populations of Triatoma infestans (Hemiptera: Reduviidae) in the Gran Chaco of Argentina

Goat production is an important economic activity for rural communities in the Gran Chaco of Argentina. Goat corrals are important for the survival of peridomestic populations of Triatoma infestans. This study evaluated the impact of modifying the traditional structure of goat corrals on T. infestans populations and goat productivity in the region of Los Llanos (La Rioja). Thirty-nine experimental corrals were constructed and 57 traditional corrals were used as controls. We evaluated the infestations of the control and experimental corrals for five years following construction of the structures. The results showed that the new structures did not prevent the colonization, although it enhanced the detection of infestation at low densities of T. infestans. No significant difference was found in T. infestans population abundance between control and experimental corrals, probably because of the different detectability in the two types of structures, especially among the small nymphs. Although goat productivity average was higher in experimental than in control corrals, no significant difference was found because of high variability. The new structures can be used as a complement to promote the development of rural communities. Acceptability and adoption of the new corrals by the owners was high, as the enclosures offered better protection for the goats, increased growth of kids and facilitated herd handling.     

Toxicity of non-pyrethroid insecticides against Triatoma infestans (Hemiptera: Reduviidae)

Triatoma infestans (Klug) is the main vector of Chagas disease, which is a public health concern in most Latin American countries. The prevention of Chagas disease is based on the chemical control of the vector using pyrethroid insecticides. In the last decade, different levels of deltamethrin resistance have been detected in certain areas of Argentina and Bolivia. Because of this, alternative non-pyrethroid insecticides from different chemical groups were evaluated against two T. infestans populations, NFS and El Malá, with the objective of finding new insecticides to control resistant insect populations. Toxicity to different insecticides was evaluated in a deltamethrin-susceptible and a deltamethrin-resistant population. Topical application of the insecticides fenitrothion and imidacloprid to first nymphs had lethal effects on both populations, producing 50% lethal dose (LD50) values that ranged from 5.2-28 ng/insect. However, amitraz, flubendiamide, ivermectin, indoxacarb and spinosad showed no insecticidal activity in first instars at the applied doses (LD50 > 200 ng/insect). Fenitrothion and imidacloprid were effective against both deltamethrin-susceptible and deltamethrin-resistant populations of T. infestans. Therefore, they may be considered alternative non-pyrethroid insecticides for the control of 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.     

Role of enhanced detoxication in a deltamethrin-resistant population of Triatoma infestans (Hemiptera, Reduviidae) from Argentina

Deltamethrin and other pyrethroids have been extensively used in Argentina since 1980, for the chemical control of Triatoma infestans Klug (Hemiptera: Reduviidae). Recently, resistance to deltamethrin was detected in field populations by the survival of bugs exposed by topical application to the diagnostic dose estimated on the CIPEIN susceptible strain. Results of the current study showed low resistant ratios (RRs) to deltamethrin for the resistant populations (RR ranged from 2.0 for San Luis colony to 7.9 for Salta colony). Biochemical studies were made on the most resistant colony (Salta) and the susceptible strain (CIPEIN), in order to establish the importance of degradative mechanisms as a cause of the detected resistance. Esterase activity was measured on 3 days old first instars through phenylthioacetate and a-naphtyl acetate activities. The results showed a significant difference in no cholinesterase esterase activity from susceptible (7.6 +/- 0,7 micro M S./i.min.) and Salta resistant colony (9.5 +/- 0.8 microM S./i.min.). Cytochrome p450 mono-oxygenase (p450) activity was measured on individual insects through ethoxycoumarine deethylase (ECOD) activity using a fluorescence microplate reader. The dependence of ECOD activity on age and body region of the nymphs, and pH and time of incubation were studied in order to optimize the measurement. As a result, comparative studies were performed on abdomens of 2 days old first instars at pH 7.2 and 4 h incubation time. ECOD activity of first nymphs was significantly lower in the susceptible colony (61.3 +/- 9.08 pg ECOD/ insect) than in the resistant one (108.1+/- 5.7 pg ECOD/ insect). These results suggest that degradative esterases (no-cholinesterase) and mono-oxygenases cytochrome p450, play an important role in the resistance to deltamethrin in Salta colony from Argentina.     

Fine structure of the salivary glands of Triatoma infestans (Hemiptera: Reduviidae)

The fine structure of the salivary glands of adult Triatoma infestans (Hemiptera: Reduviidae) bugs has been analyzed. Stereomicroscopy and scanning electron microscopy showed that each insect presents a pair of salivary glands, each pair containing three distinct units (main, supplementary, and accessory) with different sizes and colors. Transmission electron microscopy demonstrated that all gland units consist of a monolayer of epithelial cells surrounding a large central lumen. The gland units are enveloped by a thick basal lamina containing bundles of muscle cells. Microvilli are present at the apical plasma membrane domain of the gland cells, thus enlarging the available membrane area for saliva secretion towards the large gland lumen, although occasionally budding vesicles could be observed among the microvilli. Cytochemical analysis showed that the salivary gland cells of T. infestans present abundant endoplasmic reticulum profiles and several lipid droplets.     

The fine structure of the ocelli of Triatoma infestans (Hemiptera: Reduviidae)

The morphology and fine structure of the ocelli of Triatoma infestans have been analyzed by means of light and electron microscopy. The two dorsal ocelli of this species are located behind the compound eyes, looking dorsally and frontally. Externally, the ocelli are marked by the corneal lenses virtually spherical in form and limited internally by a cuticular apodeme. The lens focuses the incoming rays beyond the retina. A single layer of corneagen cells lies below the cuticular lens. The corneagen cells and photoreceptors are arranged in a cup-like fashion beneath the cuticular lens. A distal retinal zone comprises the rhabdoms, which are laterally connected in an hexagonal meshwork. A middle retinal zone comprises the receptor cell segment free of rhabdom, and a proximal zone their axons. In the middle zone, the oviform nuclei and spheroids are located. Screening pigment granules are present within the retinal cell. Spherical mitochondria are homogeneously distributed in the cytoplasm of the cell body. In the axonal zone, mitochondria are found in the peripheral region. Axons from receptor cells extend into the ocellar neuropile at the base of the ocelli, to synapse with second order neurons. The large axons of second order neurons are bundled by glial cells. The ocellar plexus exhibits a high diversity of synaptic unions (i.e. axo-dendritic, axo-axonic, dendro-axonic, and dendro-dendritic).     

Flight initiation in Triatoma infestans and T. melanosoma (Hemiptera, Reduviidae)

The flight initiation of T. infestans, the main vector of Chagas disease in the Southern Cone countries of Latin America, and of the closely-related species T. melanosoma was studied in laboratory. The results demonstrated that after the beginning of observations the peak of the flight activity was about 14 days after feeding in both species and it was usually more marked in the females than in the males, but there were no significant differences in the flight behaviour of the two species.     

Polyploidization of the BTC-32 cell line from Triatoma infestans (Hemiptera,Reduviidae)

The cytophotometric evaluation of the Feulgen-DNA content of the BTC-32 cells at passage 160 after 6 days of growth demonstrated that polyploidy is relatively frequent in this cell line. 4C values were assumed to pertain to diploid nuclei at the S or G2 phases but also to polyploid nuclei at the G1 phase. Polyploidy in 4C nuclei is assumed to be attained by endomitosis. However, there are morphological indications that polyploidization in cells with an 8C–128C Feulgen-DNA content could result from cellular and nuclear fusions, possibly mediated by viral particles present in the cell culture. Micronucleation was also frequent; it was assumed to be promoted by viral action or deficiency in some culture medium nutrient. These nuclear characteristics should be considered when using the BTC-32 cell line for monitoring the action of infective agents or their products.     

Autosomal synaptonemal complexes and sex chromosomes without axes in Triatoma infestans (Reduviidae; Hemiptera)

Meiotic and somatic cells at interphase in Triatoma infestans are characterized by the formation of a large chromocenter, which was assumed to contain the whole of the three large pairs of autosomes and the sex chromosomes. Observations with C-banding techniques show that the chromocenter is formed only by the terminal and subterminal heterochromatic blocks of the three large pairs of autosomes and the sex chromosomes. During pachytene the two largest autosomal pairs loop on themselves and their condensed ends form the chromocenter, together with the single heterochromatic end of the third autosomal pair. The X and Y chromosomes seem to associate with these condensed ends by their affinity for C-heterochromatin. During a very short pachytene stage, bivalents and synaptonemal complexes (SCs) are observed. Pachytene is followed by a very long diffuse stage, during which SCs are disassembled, multiple complexes aggregate on the inner face of the chromocenter and finally all complexes disappear and a dense material is extruded to the cytoplasm through the annuli. The 3-dimensional reconstruction of early pachytene chromocenters show 3 SCs entering and tunnelling the chromocenter, while during mid-pachytene 4 SCs enter this mass and a 5th SC is in a separate small mass. The looping of a whole SC which has both ends in the chromocenter was shown by the reconstructions. These data are interpreted as the progressive looping of the two largest bivalents during pachytene, forming finally the association of 5 bivalent ends corresponding to the 5 C-banding blocks of the large autosomal pairs. No single axis or SC that could be ascribed to the sex chromosomes was found. This agrees with the pachytene microspreads, which show only 10 SCs corresponding to the autosomal bivalents. The X and Y chromosomes are enclosed in the chromocenter, as shown by the unravelling chromocenters at diplotene-diakinesis. Thus the sex chromosomes do not form axial condensations, and this fact may be related to the ability of the X and Y chromosomes to divide equationally at metaphase I. SCsThis paper is dedicated to the memory of the late Professor Francisco A. Saez     

Microsatellite analysis of genetic structure in natural Triatoma infestans (Hemiptera: Reduviidae) populations from Argentina: its implication in assessing the effectiveness of Chagas' disease vector control programmes

The genetic structure in populations of the Chagas' disease vector Triatoma infestans was examined. Comparisons of the levels of genetic variability in populations of this species from areas with different periods since last insecticide treatment and from areas that never received treatment were also carried out. A total of 598 insects from 19 populations were typed for 10 polymorphic microsatellite loci. The average observed and expected heterozygosities ranged from 0.186 to 0.625 and from 0.173 to 0.787, respectively. Genetic drift and limited gene flow appear to have generated a substantial degree of genetic differentiation among the populations of T. infestans. Departures from Hardy-Weinberg expectations due to an excess of homozygotes suggested the presence of null alleles and population subdivision. Microgeographical analysis supports the existence of subdivision in T. infestans populations. Levels of genetic diversity in the majority of the populations of T. infestans from insecticide-treated localities were similar or higher than those detected in populations from areas without treatment. Since the populations of T. infestans are subdivided, a population bottleneck would result in independent genetic drift effects that could randomly preserve different combinations of alleles in each subpopulation. These events followed by a rapid population growth could have preserved high levels of genetic diversity. This study supports the hypothesis of vector population recovery from survivors of the insecticide-treated areas and therefore highlights the value of population genetic analyses in assessing the effectiveness of Chagas' disease vector control programmes.     

Changes in nuclear phenotype frequencies following sequential cold shocks in Triatoma infestans (Hemiptera,Reduviidae)

The nuclear phenotypes of Malpighian tubule cells in fifth instar nymphs of Triatoma infestans, one of the most important vectors of Chagas disease, were studied following sequential shocks at 0 degrees C, separated by intervals of 8 h and 24 h at 30 degrees C, under conditions of moderate fasting and full nourishment. The insects pertained to colonies reared in the laboratory and originated from domestic specimens collected in the Brazilian states of S?o Paulo (north) and Minas Gerais (south). Since nuclear phenotypes in this species are affected by single cold shocks, it was expected that these phenotypes could also be changed by sequential shocks. Nuclear phenotypes indicative of mechanisms of cell survival (nuclear fusion and heterochromatin decondensation) and cell death (apoptosis and necrosis) were observed concomitantly in all the conditions tested. Nuclear fusion and heterochromatin decondensation were not found relevant for the presumed acquisition of the cold-hardening response in T. infestans. The decreased frequency of apoptosis and necrosis following sequential cold shocks including under fasting conditions, indicated that tolerance to sequential cold shocks occurred in T. infestans of the mentioned origin.     

Isolation of Paecilomyces lilacinus (Thom) Samson (Ascomycota: Hypocreales) from the Chagas disease vector, Triatoma infestans Klug (Hemiptera: Reduviidae) in an endemic area in Argentina

A survey for entomopathogenic fungi of the Chagas disease vector Triatoma infestans was conducted in two provinces of Argentina from March–December 2003. Field-collected insects that died in the laboratory were individually maintained in moist chamber and incubated at 22 °C. Triatominae adults infected with the fungus Paecilomyces lilacinus were found at El Quebracho (27°34′S–64°31′W), Santiago del Estero province, Argentina, in December 2003. Paecilomyces lilacinus was cultured and isolated from infected insects in SDAY, PYG and MEA media. Pathogenicity tests were conducted and positive results were recorded. The median survival time (MST) of T. infestans exposed to a P. lilacinus conidial suspension was 12.8 days, and 100% mortality occurred at 30 days post-treatment. This is the first record of natural infection caused by P. lilacinus in T. infestans in the world.     

Biological effect of 1-dodecanol in teneral and post-teneral Rhodnius prolixus and Triatoma infestans (Hemiptera: Reduviidae)

Topical application of 1-dodecanol was significantly more toxic against teneral first nymphs (1-3 h old) than post-teneral first nymphs (24 h old). The lethal dose ratios were 711,500 for Rhodnius prolixus and 3613 for Triatoma infestans. No significative difference between LD50 was found when 1-dodecanol was injected in recently hatched adult R. prolixus (1-4 h old) nor in older adults (24 h old). These values were similar to those calculated for deltamethrin (an effective triatomicide), showing that 1-dodecanol had no insecticidal properties when it was applied by injection. Topical application of high dose of 1-dodecanol (1 microg/i) on teneral first nymphs of R. prolixus, produced an interruption of the darkening process of the cuticle, and probably in the development of its physiological properties.     

Ultrastructure and receptor cell responses of the antennal grooved peg sensilla of Triatoma infestans (Hemiptera: Reduviidae)

Ultrastructural examination of grooved-peg (GP) sensilla on the antenna of fifth instar Triatoma infestans nymphs by scanning electron microscopy and transmission electron microscopy reveal that they are 8–18 μm long with a diameter of about 2–2.8 μm at the non-articulated base. Some pegs have a terminal pore. These double-walled wall-pore (dw-wp) sensilla have an outer cuticular wall with 13–18 longitudinal grooves at the distal part of the peg. Groove channels are present at the bottom of the grooves from which radial spoke channels lead into the inner sensillum-lymph cavity. A dendrite sheath connects the tip of the thecogen cell to the inner cuticular wall thus forming separated outer and inner sensillum-lymph cavities. Four or five bipolar receptor cells are ensheathed successively within the GP sensilla by the thecogen cell, trichogen and tormogen cells. The inner dendritic segments of each sensory cell give rise at the ciliary constriction to an unbranched outer dendritic segment which can reach the tip of the sensillum.Electrophysiological recordings from the GP sensilla indicate that they house NH₃, short-chain carboxylic acid and short-chain aliphatic amine receptor cells and can be divided into three functional sub-types (GP 1–3). All GP sensilla carry a receptor cell excited by aliphatic amines, such as isobutylamine, a compound associated with vertebrate odour. GP type 1 and 2 sensilla house, in addition, an NH₃-excited cell whereas the type 2 sensilla also contains a short-chain carboxylic acid receptor. No cell particularly sensitive to either NH₃ or carboxylic acids was found in the grooved-peg type 3 sensilla. GP types 1, 2 and 3 represent ca. 36, 10 and 43% of the GP sensilla, respectively, whereas the remaining 11% contain receptor cells that manifest normal spontaneous activity but do not respond to any of the afore mentioned stimuli.     

Evaluation of hyperactivity produced by pyrethroid treatment on third instar nymphs of Triatoma infestans (Hemiptera: Reduviidae)

The hyperactivity produced in third instar nymphs of Triatoma infestans by their exposure to films of deltamethrin or cis-permethrin was evaluated. Both pyrethroids produced a significant increase in locomotor activity at 26 and 36°C but not at 16°C. At 26°C, only deltamethrin produced hyperactivity when topically applied on the dorsal side of the nymphs abdomen. However, both pyrethroids produced hyperactivity when topically applied on the head of the nymphs. Hyperactivity was not observed when nymphs were treated with N-ethylmaleimide (20 μg/insect) before exposure to the pyrethroids. The effect of both insecticides on locomotor activity reversed the inhibitory influence on locomotion elicited by contact with the walls of the experimental arena (thigmotaxis). Arch. Insect Biochem. Physiol. 35:323–333, 1997.© 1997 Wiley-Liss, Inc.     

Eye pigments of the blood-sucking insect, Triatoma infestans Klug (Hemiptera, Reduviidae).

The pigmentation of black (wild) and red (mutant) eyes of Triatoma infestans was studied spectrophotometrically and compared with red-eyed (wild) and white-eyed (mutant) forms of Drosophila melanogaster. The spectral absorption profiles of the black and red eye pigments of T. infestans were similar to each other and to that of the wild-type eyes of D. melanogaster. The similarity to the wild form of D. melanogaster indicated that both eye forms of T. infestans contained ommochromes of the xanthommatin type, a finding confirmed by ascending paper chromatography. Pteridines, melanins, and ommins were not detected as eye pigments in T. infestans. The eye color difference in T. infestans was assumed to be a function of the xanthommatin concentration, with a smaller content of ommochrome in red eyes, although this probably did not affect the insect's visual acuity. These data support other findings regarding the similarities between black- and red-eyed specimens of T. infestans for other characteristics.     

Triatoma sordida St?l 1859 (Hemiptera, Reduviidae: Triatominae) in palms of northeastern Argentina.

Several palms species provide an important habitat for triatomines and associate vertebrates in tropical America. The objective of this work is to identify the triatomine species living in the palms of rural areas in the Province of Corrientes, and to estimate the potential epidemiological risk they represent for the residents of nearby houses. The survey was carried out in a palm community in Colonia Laurel, Department San Roque, Province of Corrientes, Argentina. Samplings were performed in October, November and December 1998; January, February and March 1999; May and June 1999. Thirty palms: 27 (90%) Butia yatay (Mart.) Becc. and 3 Acrocomia aculeata (Jacq.) Lodd ex Mart. were dissected. Triatoma sordida St?l 1859 was found in 96.2% of B. yatay and in all the A. aculeata palms. A total of 272 live T. sordida was collected; 36 of them (13.2%) were found in bird nests in the frond and the remainder in other locations of the tree. The mean number of triatomines per palm was 9.6 (range 1-60, mode 2). T. sordida was collected during all the sampling months and all stages were present at all seasons. The highest population density was reached in spring and the lowest in autumn. Trypanosoma cruzi was detected in 38.5% in feces of 174 examined insects and identified as such, both by microscopical examination and PCR. This is the first finding of T. sordida populations in B. yatay, an endemic palm of South America distributed in southern Brazil, Uruguay and northeastern Argentina. The high infection prevalence found in this work suggests that T. sordida plays an essential role in the maintenance of the wild T. cruzi transmission cycle in northeastern Argentina.     

Identification and characterization of a novel factor XIIa inhibitor in the hematophagous insect, Triatoma infestans (Hemiptera: Reduviidae)

Recently, we have cloned several Kazal-type serine protease inhibitors from the midgut of the Triatoma infestans bug. A single gene composed of multi Kazal-type domains, in tandem, encodes these inhibitors. In this work, we describe the purification and characterization of recombinant infestins 3-4 and 4, which are potent factor XIIa inhibitors (KI=67 pM and 128 pM, respectively). We also identified the first native factor XIIa inhibitor from a hematophagous insect. The factor XIIa inhibitory activity of infestin 4 demonstrates extremely efficient anticoagulant activity, prolonging activated partial thromboplastin time by approximately 3 times. Our results suggest that infestins perform a very important role in the T. infestans midgut during meal acquisition and digestion by controlling blood coagulation by means of inhibiting thrombin and factor XIIa.     

Genetic diversity of Triatoma infestans (Hemiptera: Reduviidae) in Chuquisaca, Bolivia based on the mitochondrial cytochrome b gene

Partial cytochrome b DNA sequences for 62 Triatoma infestans were analyzed to determine the degree of genetic variation present in populations of this insect in the northwest region of Chuquisaca, Bolivia. A total of seven haplotypes were detected in the localities sampled. The phylogenetic relationship and population genetic structure of the haplotypes found in this region, indicate that there is greater variation in this relatively small region of Bolivia than what has been previously reported by studies using the same gene fragment, for more distant geographic areas of this country. In addition, a comparison of rural and peri-urban localities, indicate that there is no difference in the genetic variation of T. infestans between these two environments.