Intraspecific and Interspecific Phenotypic Differences Confirm the Absence of Cryptic Speciation in Triatoma sordida (Hemiptera, Triatominae).

Triatoma sordida is an endemic Chagas disease vector in South America, distributed in Brazil, Bolivia, Paraguay, and Uruguay. Chromosomal, molecular, isoenzymatic, and cuticular hydrocarbon pattern studies indicate cryptic speciation in T. sordida. Recently, T. rosai was described from specimens from Argentina initially characterized as T. sordida. Although several authors assume that the speciation process that supports this differentiation in T. sordida is the result of cryptic speciation, further morphological and/or morphometric studies are necessary to prove the application of this evolutionary event, because the only morphological intraspecific comparison performed in T. sordida is based on geometric morphometry and the only interspecific comparison made is between T. rosai and T. sordida from Brazil that evaluated morphological and morphometric differences. Based on this, morphological analyses of thorax and abdomen using Scanning Electron Microscopy and morphometric analyses of the head, thorax, and abdomen among T. sordida from Brazil, Bolivia, and Paraguay, as well as T. rosai, were performed to assess whether the evolutionary process responsible for variations is the cryptic speciation phenomenon. Morphological differences in the thorax and female external genitalia, as well as morphometric differences in the head, thorax, abdomen, pronotum, and scutellum structures, were observed. Based on this, the evolutionary process that supports, so far, these divergences observed for T. sordida populations/T. sordida subcomplex is not cryptic speciation. Moreover, we draw attention to the necessity for morphological/morphometric studies to correctly apply the cryptic species/speciation terms in triatomines.

Morphological and Chromatic Variation in Four Populations of Triatoma mexicana (Hemiptera: Reduviidae).

Triatoma mexicana is an endemic species of Mexico and is distributed in the states of Hidalgo, Queretaro, Guanajuato, and San Luis Potosi, being naturally infected with Trypanosoma cruzi, which increases its importance in the region. The species description was made in 1848, but there are only a few studies on its morphology, biology, and behavior. The present manuscript shows the presence of morphological and chromatic variations among populations of T. mexicana from the states of Hidalgo (Valle del Mezquital and Meztitlan), Guanajuato and Queretaro. The study employed 136 specimens collected in four locations. Scanning electron microscopy was used to study the morphological characteristics of the head, pronotum, and scutellum; also, we measured the width of the abdomen and the total length in the specimens of each population. The morphometric analysis considered 19 variables in the previous structures. Significant differences were found in the dimensions of the head and pronotum, but not in the scutellum; there is clear discrimination among the four proposed populations. The chromatic patterns observed in the connexivum go from yellow to brown and show some significant differences related to geographical origin. The set of evaluated characters showed a higher degree of difference in the population of Guanajuato, clearly separating from the rest of the populations, indicating the possibility of a divergence process. The characteristics observed in the remaining populations could be adaptive responses to their habitat.

Structure and function of the stylets of hematophagous Triatominae (Hemiptera: Reduviidae), with special reference to Dipetalogaster maxima.

Kissing bugs (Hemiptera: Reduviidae: Triatominae) are able to bend their rod-like maxillae while searching for blood vessels in the tissue of their vertebrate hosts. Little is known about the working mechanisms of these bending movements and the distal opening of the food channel. We compared the morphological structure of the stylets (mandibles and maxillae) of four triatomine species and analyzed the feeding process of Dipetalogaster maxima (Uhler, 1894). The maxillae of triatomine bugs are interlocked by a tongue-and-groove system, allowing longitudinal sliding. While penetrating the host tissue, the animals perform rapid alternate back and forth movements of the maxillae. The resistance of the surrounding tissue pushes the asymmetric apex of the maxillae away from its straight path, i.e., if one individual maxilla is protracted alone, its tip curves inwards, and the other maxilla follows. Once a blood vessel is tapped, the spine-like tip of the left maxilla splays outwards. Apically, each of the maxillae features an abutment, the left one exhibiting a notch that presumably facilitates splaying. The mechanical interaction of the two maxillary abutments enables the distal opening of the food channel but might also support the movements of the maxillary bundle attributable to different bending moment distributions.

Development of conventional and real-time multiplex PCR-based assays for estimation of natural infection rates and Trypanosoma cruzi load in triatomine vectors.

BACKGROUND: Chagas disease is a complex anthropozoonosis with distinct domestic and sylvatic mammal species acting as potential reservoirs. The diversity of vector species and their habitats are among the factors that hinder the control of the disease. Control programs periodically monitor the prevalence of T. cruzi infection in insect bugs through microscopical observation of diluted feces. However, microscopy presents limited sensitivity in samples with low parasite numbers, difficulties in examining all evolutionary stages of the insect and may in turn be limited to differentiate T. cruzi from other morphologically similar trypanosomatids. Here, we report two highly sensitive and accurate methodologies to infer T. cruzi infection rates and to quantify parasite load in the gut of field-collected triatomines. METHODS: Triatomines were manually collected in the period 2011-2012 and 2014-2015, in domestic, peridomestic or sylvatic habitats in rural areas of 26 municipalities, encompassing three distinct Brazilian biomes: Caatinga, Cerrado and Atlantic Rainforest. Following morphological and taxonomical identification, the search for flagellated protozoa was performed by optical microscopy. A conventional PCR targeting T. cruzi kDNA and a TaqMan qPCR directed to the parasite nuclear satellite DNA (SAT) were developed, both in multiplex, with the triatomine 12S subunit ribosomal RNA gene, used as internal amplification control. Both methods were used for detection (kDNA-PCR) and parasite load quantification (SAT-DNA-qPCR), to investigate T. cruzi infection in captured triatomines. RESULTS: The combined methods were assayed on a panel of 205 field-collected triatomine samples. Diagnostic analysis revealed 21% positivity for the kDNA-PCR, whereas microscopic examination enabled identification of T. cruzi in only 7.0% of the PCR-positive samples. Negative PCR results were confirmed by the absence of T. cruzi flagellates using microscopy. Caatinga biome yielded the highest T. cruzi infection rate (60%), followed by the Atlantic Rainforest and Cerrado with 7.1 and 6.1%, respectively. In addition, a wide range distribution of parasite load, varying from 8.05 × 10-2 to 6.31 × 1010 was observed with a median of 2.29 × 103 T. cruzi/intestine units. When parasite load was analyzed by triatomine species, a significantly higher median was found for Panstrongylus lutzi in comparison with Triatoma brasiliensis. CONCLUSIONS: Our results demonstrate highly sensitive PCR-based methodologies to monitor T. cruzi infection in triatomines. In addition, the qPCR assay offers the possibility of further evaluation parasite load, as a promising biomarker of the vectorial capacity of triatomines in Chagas disease endemic areas.

Sperm dimorphism in the Triatoma brasiliensis species complex and its applications.

Morphological and structural features of the sperm of the Triatoma brasiliensis Neiva, 1911 (Hemiptera: Reduviidae) species complex were examined in this first study investigating the sperm of Heteroptera and the genus Triatoma. Males were dissected and their seminal vesicles removed. For measurement, seminal vesicles were squashed on glass slides, spread, fixed and observed under a photomicroscope. The images were analysed and measures of sperm were made. Data were analysed using one-way analysis of variance and Tukey's test to detect differentiation among taxa. Furthermore, seminal vesicles were prepared for studies of transmission electron microscopy. All taxa studied showed polymorphic (short and long) sperm. The sperm of Triatoma brasiliensis macromelasoma was significantly longer (in total length) than that of the other four members of the complex, which supports the hypothesis of hybrid speciation of this member of the complex as an increase in the size of typical hybrids under heterosis was previously shown. The sperm cells of the five taxa have similar ultrastructural morphology. The ultrastructural features observed confirm the hypothesis, raised by previous studies, that they are synapomorphic to the suborder Heteroptera.

Revalidation of Triatoma bahiensis Sherlock & Serafim, 1967 (Hemiptera: Reduviidae) and phylogeny of the T. brasiliensis species complex.

Triatoma bahiensis Sherlock & Serafim, 1967, T. lenti Sherlock & Serafim, 1967, and T. pessoai Sherlock & Serafim, 1967 were described based on material collected in the Brazilian state of Bahia. These species were later included in the T. brasiliensis complex based on their geographic distribution. Triatoma bahiensis and T. pessoai were subsequently synonymized with T. lenti. However, the phylogenetic position of T. lenti within the T. brasiliensis complex has remained doubtful. This study aims to assess the taxonomic status of T. bahiensis and to infer the phylogenetic relationships between T. lenti, T. bahiensis and the other members of the T. brasiliensis species complex. The identities of the species in concern were confirmed by comparisons with high resolution photos of the respective type materials; lectotypes are designated for T. pessoai and T. bahiensis. Morphological, morphometric, molecular, and cytogenetic approaches as well as experimental crosses were used. The low viability of experimental crosses combined with morphological and morphometric data allow the differentiation of T. bahiensis and T. lenti. Pairwise cyt b sequence divergence between T. lenti and T. bahiensis was 2.5%. Cytogenetic and molecular analyses grouped T. lenti and T. bahiensis as members of the T. brasiliensis complex. These results revalidate the specific status of T. bahiensis.

Biological, morphological and morphometric studies of Triatoma melanocephala Neiva & Pinto, 1923 (Hemiptera: Reduviidae: Triatominae).

Triatoma melanocephala Neiva & Pinto is found in the Brazilian states of Bahia, Paraíba, Pernambuco, Rio Grande do Norte, and Sergipe. In addition to the species' specific description, eight other articles on this insect were found in the literature. In this study, data was obtained on the morphology, morphometry, and life cycle of T. melanocephala, since this vector is of epidemiological and taxonomic importance. The specimens studied were obtained from a colony that has been kept at the Triatomine Insectarium of the College of Pharmaceutical Sciences of São Paulo State University's in Brazil. The morphological studies were performed using scanning electron microscopy. These studies characterized the eggs, the external adult female genitalia, and the ninth ventral abdominal segments of male and female nymphs. The morphometric studies characterized the five nymphal instars and the adult stage by measuring the head, thorax, abdomen, antennae, and mouthparts parameters. The life cycle of T. melanocephala was developed starting by 15 couples in the fifth instar. They were fed on Swiss mice every two weeks and observed daily. During daily observation, minimum temperature, maximum temperature, and relative humidity of the laboratory were measured. The results of the biological, morphometric, and morphological studies have increased the knowledge available on T. melanocephala.

Immunofluorescence and ultrastructural analysis of the chromatoid body during spermatogenesis of Triatoma platensis and T. rubrovaria (Hemiptera, Triatominae).

This study sought to analyze spermatogenesis in two species of triatomines (Triatoma rubrovaria and T. platensis) by focusing on the chromatoid body (CB) during three stages of spermatogenesis (spermatocytogenesis, meiosis, and spermiogenesis). The cytochemistry technique known as silver impregnation revealed nucleolar persistence. We suggest that this phenomenon is fundamental to the formation of the CB during spermatogenesis, as it allows for the nucleolus or nucleolar fragments to maintain their transcriptional activity during the entire meiosis phase and to apply all transcribed RNA to CB formation. The ultrastructural analysis of T. platensis and T. rubrovaria spermatids revealed the presence of the nucleolus within the spermatid nucleus, as well as the CB near the nuclear membrane. Immunofluorescence for fibrillarin revealed the presence of protein in both the nucleolus and the cytoplasm of spermatogonia. Based on these findings, we suggest that the formation of the CB begins during the first phase of spermatogenesis, or spermatocytogenesis. Furthermore, we also observed the presence of fibrillarin protein in the CB near the elongating spermatids. Unlike the spermatogonia, spermatids showed no fibrillarin markings in the nucleolar region, a finding which is consistent with the lack of post-meiotic transcriptional activity during triatomine spermiogenesis. Thus, this study suggests that the formation of the CB begins during spermatocytogenesis and is intensified by transcriptional activity when nucleolar persistence occurs in meiosis. Moreover, the findings are consistent with the absence of transcriptional activity in the nucleolus during spermiogenesis, and they demonstrate that all transcriptional activity during spermatid differentiation is supported by the CB.

Development and glycoprotein composition of the perimicrovillar membrane in Triatoma (Meccus) pallidipennis (Hemiptera: Reduviidae).

Hemipterans and thysanopterans (Paneoptera: Condylognatha) differ from other insects by having an intestinal perimicrovillar membrane (PMM) which extends from the base of the microvilli to the intestinal lumen. The development and composition of the PMM in hematophagous Reduviidae depend on factors related to diet. The PMM may also allow the human parasite Trypanosoma cruzi, the etiological agent of human Chagas Disease, to establish and develop in this insect vector. We studied the PMM development in the Mexican vector of Chagas Disease, Triatoma (Meccus) pallidipennis. We describe changes in the midgut epithelial cells of insects in response to starvation, and at different times (10, 15 and 20 days) after bloodfeeding. In starved insects, the midguts showed epithelial cells closely connected to each other but apparently free of PMM with some regions being periodic acid-Schiff (PAS-Schiff) positive. In contrast, the PMM was evident and fully developed in the midgut region of insects 15 days after feeding. After this time, the PMM completely covered the microvilli and reached the midgut lumen. At 15 days following feeding the labeled PAS-Schiff increased in the epithelial apex, suggesting an increase in carbohydrates. Lectins as histochemical reagents show the presence of a variety of glycoconjugates including mannose, glucose, galactosamine, N-acetyl-galactosamine. Also present were N-acetyl-glucosamine and sialic acid which contribute to the successful establishment and replication or T. cruzi in its insect vectors. By means of scanning electron microscopy (SEM) and transmission electron microscopy (TEM), the formation and structure of the PMM is confirmed at 15 days post feeding. Our results confirmed the importance of the feeding processes in the formation of the PMM and showed the nature of the biochemical composition of the vectors' intestine in this important Mexican vector of Chagas disease.

Morphological study of eggs from five Mexican species and two morphotypes in the genus Triatoma (Laporte, 1832).

We describe and compare the morphology and morphometry of the egg exochorion for five species and two morphotypes of Mexican triatomines with scanning electron microscopy. The results show differences in egg ornamentation for each species, including between morphotypes. Polygonal ornamentation was observed in each species, including pentagons, octagons and, in certain cases, a majority of hexagons. We observed small perforations in T. protracta protracta, small spheres on the T. lecticularia polygons, a crown with festoons in T. barberi, and less complex ornamentation in T. mexicana. Through morphometric analysis, significant differences in egg dimensions were determined for the studied species.

Ultrastructure and morphometry of eggs of Triatoma rubrovaria (Blanchard, 1843), Triatoma carcavalloi Juberg, Rocha & Lent, 1998 and Triatoma circummaculata (Stål, 1859) (Hemiptera-Reduviidae-Triatominae).

This study analyzed the body and the operculum of eggs of Triatoma rubrovaria, T. carcavalloi and T. circummaculata, considered sylvatic species that live in sympatry.Triatoma rubrovaria is currently considered the most important vector of Trypanosoma cruzi in the rural areas of the state of Rio Grande do Sul, followed by T.circummaculata. Significant differences other than morphometry have been observed in the egg structures of the three species using traditional microscopy and scanning electron microscopy. Triatoma circummaculata eggs are smaller than those of T.rubrovaria and T. carcavalloi. The average number of perforations in corionic cells in the egg body is higher for the T. rubrovaria. The average number of perforations in the operculum cell is higher in T. circummaculata. This is the first morpho-structural description of T. carcavalloi eggs. These results widen the concept of these three species and create new subsidies for the entomological monitoring in areas in which these vectors may infest human living quarters.

Cytogenetic characterization reveals differences in nuclear organization of cystic cells among Brazilian species of Triatoma (Heteroptera, Reduviidae).

Cytogenetic studies in triatomines have described the occurrence of holokinetic chromosomes, heterochromatin distribution and the location of rDNA (ribosomal DNA) sites, but few aspects of nuclear organization in this group have been discussed. We have focused on ultrastructural and cytogenetic features and differences in cystic cells of seminiferous tubules between five species of Triatoma. Cystic cells showed evidence of polyploidy events and heterochromatic blocks appeared predominantly in the central region of the nuclei. Cytogenetic analyses showed that there was variation in chromocenter number between species, and that the central regions were AT-rich [DAPI+ (4',6-diamidino-2-phenylindole+)], whereas the periphery was CG-rich (CMA+). Another characteristic was the distribution of 45S rDNA, which differed according to the chromosomal location of this sequence. In all we have compared aspects of nuclear organization, polyploidy, heterochromatin, rDNA site distribution and methylation levels, as well as the relationships between five species of Triatoma from a cystic cell perspective.

The morphological variation of the eggs and genital plates of two morphotypes of Triatoma protracta Uhler, 1894.

The control of triatomine insects is necessary because these insects are the principal vectors of Trypanosoma cruzi, the agent of Chagas disease. Nevertheless, some of these vectors, such as Triatoma protracta, have not been studied adequately and their importance and taxonomic status has not yet been defined in detail and must be reevaluated in view of the continuing taxonomic uncertainties associated with the species. To help clarify the taxonomic status of T. protracta, the eggs and genital plates of two morphotypes were analyzed. Qualitative and quantitative morphological differences were observed in two morphotypes, designated T. p. protracta and T. p. nahuatlae according to Ryckman (1962). The morphotype T. p. protracta exhibited large and wide eggs with pores forming large padded polygonal structures, whereas the eggs of the morphotype T. p. nahuatlae were small and smooth. The size of the 9(th) genital urosternite was longer and wider in females in contrast to males in both morphotypes. However, these size differences were relatively greater in T. p. protracta. The high morphological variation found between the morphotypes of T. protracta suggests that they should be separated. Accordingly, it is probable that this group should be re-classified.

Chromatoid body: remnants of nucleolar proteins during spermatogenesis in triatomine (Heteroptera, Triatominae).

In this study, we analyzed fibrillarin nucleolar protein expression in CBs of spermatogenic cells from testicular follicles of Triatoma sordida and Triatoma infestans. In the structural and ultrastructural analysis, it was used impregnation by silver ions, immunocytochemistry, immunofluorescence and transmission electron microscopy using antibodies against fibrillarin. Regarding the results, the fibrillarin nucleolar protein marked the nucleus and some cytoplasmic spots of germ cells during spermatogenesis in triatomines. These data suggest that fibrillarin could be a constituent of the CB that was most likely derived from nucleolar fragmentation. This is the first time that fibrillarin protein expression has been shown in the CB during spermatogenesis progression in triatomines. The knowledge regarding CB constituents may help to expand the understanding of the physiological role of this structure and the role that it plays in the reproductive biology of triatomines, which are vectors of Chagas Disease.

Ultrastructural localization of basic proteins and carbohydrates in male accessory glands of two Triatoma species (Hemiptera, Reduviidae, Triatominae).

The male accessory glands in Triatoma are tubular and produce substances with some functions related to production of the spermatophore. In the current study, the cytochemistry of male accessory glands was evaluated in starved Triatoma brasiliensis and adult Triatoma melanica. The storage of carbohydrates and proteins in T. melanica male accessory glands occurs earlier than in T. brasiliensis. In addition, the occurrence of eletron-lucent granules without carbohydrates and proteins suggests that other substances are released by these glands, which may be used for lubrication of the male genitalia. Male T. brasiliensis has more intense secretory activity in the fifth day of adult life, which may indicate a higher reproductive capacity. The analysis of lipid production in male accessory glands can contribute to the knowledge of spermatophore formation in these species.

Capsid protein identification and analysis of mature Triatoma virus (TrV) virions and naturally occurring empty particles.

Triatoma virus (TrV) is a non-enveloped +ssRNA virus belonging to the insect virus family Dicistroviridae. Mass spectrometry (MS) and gel electrophoresis were used to detect the previously elusive capsid protein VP4. Its cleavage sites were established by sequencing the N-terminus of the protein precursor and MS, and its stoichiometry with respect to the other major capsid proteins (VP1-3) was found to be 1:1. We also characterized the polypeptides comprising the naturally occurring non-infectious empty capsids, i.e., RNA-free TrV particles. The empty particles were composed of VP0-VP3 plus at least seven additional polypeptides, which were identified as products of the capsid precursor polyprotein. We conclude that VP4 protein appears as a product of RNA encapsidation, and that defective processing of capsid proteins precludes genome encapsidation.

Estudo morfológico e ultraestrutural da glândula salivar de Rhodnius prolixus, Panstrongylus megistus e Triatoma infestans (Hemiptera, Triatominae)

Os triatomíneos são vetores do Trypanosoma cruzi e do Trypanosoma rangeli. Apresentam hábito hematófago durante todo o ciclo de vida. Durante o repasto, secretam uma saliva que antagoniza os sistemas hemostático, inflamatório e imunológico do hospedeiro vertebrado. A fim de caracterizar os aspectos morfológicos da glândula salivar, foi desenvolvido um estudo do crescimento desse órgão em triatomíneos de três espécies: Rhodnius prolixus, Panstrongylus megistus e Triatoma infestans. Os insetos foram cedidos pelo insetário do Laboratório de Triatomíneos e Epidemiologia da Doença de Chagas (CPqRR - FIOCRUZ). As glândulas salivares foram dissecadas e processadas para microscopia de luz, de fluorescência, laser confocal e eletrônica de varredura. Os resultados indicaram que a glândula é morfologicamente semelhante durante todo o ciclo de vida do inseto, sendo formada por um par de lobos, cada qual formado por dois (R. prolixus) ou três lóbulos (P. megistus e T. infestans). Durante o ciclo de vida, a glândula quadruplicou o seu tamanho inicial, devido ao crescimento celular, visto que o número de células permaneceu constante.

Os lóbulos se mostraram formados por um epitélio simples que delimita o lúmen (onde a saliva é armazenada), revestidos por fibras musculares (cuja contração permite a liberação da saliva), traquéias e nervos. Serotonina apresentou intensa atividade na glândula das três espécies analisadas durante alimentação. Em R. prolixus, atividade dopaminérgia e de tirosina-hidroxilase (enzima da via das catecolaminas) também foram observadas. Dobras de membrana foram visualizadas na região apical das células e associadas ao aumento da área de secreção (tipo merócrina). Além disso, a presença de vesículas de secreção e células inteiras (estas observadas apenas em R. prolixus) no lúmen indicaram secreções do tipo apócrina e holócrina, respectivamente. Bactérias Gram-negativas e Gram-positivas foram observadas dentro das células e do lúmen do lóbulo posterior de T. infestans

Estudo morfológico e ultraestrutural da glândula salivar de Rhodnius prolixus, Panstrongylus megistus e Triatoma infestans (Hemiptera, Triatominae) / Morphological and ultrastructural study of salivary gland of Rhodnius prolixus and Triatoma infestans Panstrongylus megistus (Hemiptera, Triatominae)

Os triatomíneos são vetores do Trypanosoma cruzi e do Trypanosoma rangeli. Apresentam hábito hematófago durante todo o ciclo de vida. Durante o repasto, secretam uma saliva que antagoniza os sistemas hemostático, inflamatório e imunológico do hospedeiro vertebrado. A fim de caracterizar os aspectos morfológicos da glândula salivar, foi desenvolvido um estudo do crescimento desse órgão em triatomíneos de três espécies: Rhodnius prolixus, Panstrongylus megistus e Triatoma infestans. Os insetos foram cedidos pelo insetário do Laboratório de Triatomíneos e Epidemiologia da Doença de Chagas (CPqRR - FIOCRUZ). As glândulas salivares foram dissecadas e processadas para microscopia de luz, de fluorescência, laser confocal e eletrônica de varredura. Os resultados indicaram que a glândula é morfologicamente semelhante durante todo o ciclo de vida do inseto, sendo formada por um par de lobos, cada qual formado por dois (R. prolixus) ou três lóbulos (P. megistus e T. infestans). Durante o ciclo de vida, a glândula quadruplicou o seu tamanho inicial, devido ao crescimento celular, visto que o número de células permaneceu constante.

Os lóbulos se mostraram formados por um epitélio simples que delimita o lúmen (onde a saliva é armazenada), revestidos por fibras musculares (cuja contração permite a liberação da saliva), traquéias e nervos. Serotonina apresentou intensa atividade na glândula das três espécies analisadas durante alimentação. Em R. prolixus, atividade dopaminérgia e de tirosina-hidroxilase (enzima da via das catecolaminas) também foram observadas. Dobras de membrana foram visualizadas na região apical das células e associadas ao aumento da área de secreção (tipo merócrina). Além disso, a presença de vesículas de secreção e células inteiras (estas observadas apenas em R. prolixus) no lúmen indicaram secreções do tipo apócrina e holócrina, respectivamente. Bactérias Gram-negativas e Gram-positivas foram observadas dentro das células e do lúmen do lóbulo posterior de T. infestans

Ultrastructural analysis of the nucleolar aspects at spermiogenesis in triatomines (Heteroptera, Triatominae).

In this study the ultrastructural technique was used to analyze seminiferous tubule cells of the triatomine species Panstrongylus megistus, Rhodnius pallescens and Triatoma infestans. The data obtained provided evidence of the phenomenon known as persistence of the nucleolar material in initial spermatids at early differentiation. Our results confirmed the presence of the nucleolus and its products during spermiogenesis up to the formation of the axoneme and during spermatid elongation in all three species studied, similar to the process that takes place during cell division. In early spermatids, the nucleoli had a reticulate appearance and a well defined nucleolonema in P. megistus; showed a clear distinction between the fibrillar and the granular component in T. infestans; and had a compact aspect in R. pallescens. In this study, ultrastructural analyses at spermiogenesis indicated that these nucleolar products may represent RNP complexes that will probably be needed at early spermiogenesis when important changes such as chromatin condensation and acrosome and flagellum formation take place. Therefore, it was concluded from the ultrastructural analysis that the triatomine nucleolus does not totally disappear but remains as corpuscles that gather to form the next nucleolar cycle that in the case of meiosis, will be completed if fertilization occurs and a zygote is formed.

Characterization of the external female genitalia of six species of Triatominae (Hemiptera: Reduviidade) by scanning electron microscopy.

By macroscopic and microscopic dorsal side observation, it was noted that the IX and X segments of two species each of Panstrongylus and Triatoma terminate in an elongated way, whereas they terminate abruptly in the two species of Rhodnius. Scanning observation of the dorsal, ventral, lateral and posterior sides of the female genitalia of Panstrongylus herreri, Panstrongylus megistus, Rhodnius colombiensis, Rhodnius prolixus, Triatoma infestans and Triatoma vitticeps revealed that these six species are generally and specifically distinguished based on these elements. We describe several components that distinguish P. herreri from P. megistus: four on the dorsal side: the VII, VIII, IX and X segments, on the ventral view, three: VII sternite, VIII gonocoxite and VIII gonapophyse, on the lateral view one character, VIII gonocoxite and on the posterior view three characters: VIII and IX gonocoxite and XI gonopophyse. Comparing R. colombiensis and R. prolixus, there were three distinct characters on the dorsal side: the VII, VIII and X segments, on the ventral view three characters: the IX and X segments and VIII gonocoxite and on the posterior view four characters: the VIII, IX, X segments and VIII gonapophyse that distinguish the two species. T. infestans and T. vitticeps have four different characters on the dorsal side: the VII, VIII, IX and X segments, on the ventral view four characters: the VII and X segments, VIII gonocoxite and VIII gonapophyse, on the lateral view two characters, IX and X segments and on the posterior view four characters: the IX and X segments, VIII gonocoxite and VIII gonapophyse that distinguish the two species. Examination of the external female genitalia of six triatomine species by scanning suggests that these components are useful for taxonomical studies.