Karyotype Evolution in Triatominae (Hemiptera, Reduviidae): The Role of Chromosomal Rearrangements in the Diversification of Chagas Disease Vectors.

Several cytogenetic studies have already been performed in Triatominae, such that different karyotypes could be characterized (ranging from 2n = 21 to 25 chromosomes), being the changes in the number of chromosomes related mainly to fusion and fission events. These changes have been associated with reproductive isolation and speciation events in other insect groups. Thus, we evaluated whether different karyotypes could act in the reproductive isolation of triatomines and we analyzed how the events of karyotypic evolution occurred along the diversification of these vectors. For this, experimental crosses were carried out between triatomine species with different karyotypes. Furthermore, based on a phylogeny with 88 triatomine taxa (developed with different molecular markers), a reconstruction of ancestral karyotypes and of anagenetic and cladogenetic events related to karyotypic alterations was performed through the ChromoSSE chromosomal evolution model. All crosses performed did not result in hybrids (prezygotic isolation in both directions). Our modeling results suggest that during Triatominae diversification, at least nine cladogenetic events may be associated with karyotype change. Thus, we emphasize that these alterations in the number of chromosomes can act as a prezygotic barrier in Triatominae (karyotypic isolation), being important evolutionary events during the diversification of the species of Chagas disease vectors.

Patterns of genome size variation in caridean shrimps: new estimates for non-gambarelloides Synalpheus species.

Genome size (GS) or DNA nuclear content is considered a useful index for making inferences about evolutionary models and life history in animals, including taxonomic, biogeographical, and ecological scenarios. However, patterns of GS variation and their causes in crustaceans are still poorly understood. This study aimed to describe the GS of five Neotropical Synalpheus non-gambarelloides shrimps (S. apioceros, S. minus, S. brevicarpus, S. fritzmueller, and S. scaphoceris) and compare the C-values of all Caridea infraorder in terms of geography and phylogenetics. All animals were sampled in the coast of São Paulo State, Brazil, and GS was assessed by flow cytometry analysis (FCA). The C-values ranged from 7.89 pg in S. apioceros to 12.24 pg in S. scaphoceris. Caridean shrimps had higher GS than other Decapoda crustaceans. The results reveal a tendency of obtaining larger genomes in species with direct development in Synalpheus shrimps. In addition, a tendency of positive biogeographical (latitudinal) correlation with Caridea infraorder was also observed. This study provides remarkable and new protocol for FCA (using gating strategy for the analysis), which led to the discovery of new information regarding GS of caridean shrimps, especially for Neotropical Synalpheus, which represents the second-largest group in the Caridea infraorder.

Biology of Chagas disease vectors: biological cycle and emergence rates of Rhodnius marabaensis Souza et al., 2016 (Hemiptera, Reduviidae, Triatominae) under laboratory conditions.

In Latin America, Chagas disease has been mostly transmitted to humans by contact with the feces or urine of triatomine species infected with the protozoan Trypanosoma cruzi. There are currently 156 species in the subfamily Triatominae, distributed in 18 genera and five tribes. The prolixus group of the genus Rhodnius is composed of 11 species. Rhodnius marabaensis was the last species described and considered in this grouping of vectors. Knowledge about the biology, ecology, and behavior of these vectors is of great epidemiological importance, and in order to expand the knowledge of the biology of R. marabaensis, this paper describes the biological cycle and emergence rates of the species under laboratory conditions. The experiment was carried out at temperatures ranging from 15.5 to 29 °C (average of 24 °C) and humidity ranging from 51.4 to 72.2 (average of 63). For each of the fifteen couples, the egg emergence rate was calculated throughout the oviposition period. The oviposition period lasted from February to September, and the emergence rate varied between 13.9 and 53.3%. R. marabaensis presented an emergence rate of 46.7% and a total biological cycle of 193 days (the mean time required for emergence (25.1 days), 1st nymphal instar (19.4 days), 2nd nymphal instar (22.1 days), 3rd nymphal instar (26.2 days), 4th nymphal instar (29.3 days), and 5th nymphal instar (70.9 days)). Based on the biological cycle of R. marabaensis and 14 other Rhodnius species already described in the literature, it was also possible to calculate the averages for the groups prolixus, pictipes, and pallescens and, mainly, for the genus Rhodnius, contributing to the knowledge of this important group of Chagas disease vectors.

Parasite × vector relationship in Chagas disease: does Trypanosoma cruzi (Chagas, 1909) infection affect the spermatogenesis of Triatoma infestans (Klug, 1834)?

The parasite-vector interaction of Chagas disease is still poorly understood and the understanding of this relationship can help in the development of new strategies to control Trypanosoma cruzi transmission, which is the etiological agent of this disease. Considering the need to know if T. cruzi can cause some pathology in the reproductive system of the Chagas disease vectors, we investigated the spermatogenesis of Triatoma infestans infected by T. cruzi through histological and cytogenetic analysis. Trypanosoma cruzi Bolivia strain infection was not pathogenic for the reproductive system of T. infestans, because all the analyzed males had normal spermatogenesis, with all phases (spermatocytogenesis, meiosis and spermiogenesis) happening without any change. Thus, we demonstrated that the presence of T. cruzi Bolivia strain does not have influence in the spermatogenesis of T. infestans and we suggest that the influences on reproductive system observed for other species were a result of the action of the parasite on gametogenesis of females.

Nucleolar persistence during spermatogenesis of the genus Rhodnius (Hemiptera, Triatominae).

The Triatominae subfamily is comprised of 18 genera and six tribes. The tribe Rhodniini is comprised of two genera (Rhodnius and Psammolestes). Nucleolar persistence is defined by the presence of the nucleolus or nucleolar corpuscles during the meiotic metaphase. To date, this phenomenon has been described for 13 species of triatomine that are included in the genera Triatoma, Rhodnius, and Panstrongylus. Thus, because the phenomenon of nucleolar persistence has been described in only two species of the genus Rhodnius, we have analyzed the nucleolar behavior during spermatogenesis of eight species of the genus Rhodnius (R. colombiensis, R. montenegrensis, R. nasutus, R. neglectus, R. neivai, R. pictipes, R. prolixus, and R. robustus), with a focus on nucleolar persistence. By means of cytogenetic analysis with silver ions, nucleolar behavior during spermatogenesis is described in the eight species of Rhodnius analyzed. In all of them nucleolar behavior was similar and the phenomenon of nucleolar persistence was often observed. Therefore, we confirm nucleolar persistence as a peculiarity of the genus Rhodnius. However, it is emphasized that new cytogenetic analysis should be performed in the Triatominae subfamily, more specifically among the 15 genera that do not exhibit the nucleolar behavior described, in order to assess whether this phenomenon is truly a synapomorphy of these hematophagous insects.

Analysis of the maternal inheritance hypothesis of the exochorium in eggs from hybrids of Chagas disease vectors.

Morphological studies applied to the taxonomy of the Triatominae cover various structures (head, wing, thorax, genitalia, and eggs). Exochorial structures of hybrid eggs were characterized and compared with the parents, demonstrating that hybrids presented characteristics identical to the exochorial pattern observed in the females of the crosses, which resulted in the hypothesis that the pattern of triatomine eggs is possibly a characteristic inherited from females. Thus, we characterized the exochorium of the eggs of several triatomine hybrids and compared them with the parents, to assess the pattern of segregation and test the hypothesis of maternal inheritance. Hybrids were obtained in at least one direction from all crosses. The analysis of the exochorium of the eggs of the hybrids showed different patterns of segregation: "exclusively paternal", "predominantly maternal", "predominantly paternal", "mutual", and "differential". Curiously, none of the hybrids evaluated presented characteristics that segregated exclusively from the female parental species. Thus, we demonstrate that the hypothesis of maternal inheritance of the exochorium pattern of eggs is not valid and we emphasize the importance of alternative/combined tools (such as integrative taxonomy) for the correct identification of these insect vectors (mainly in view of possible natural hybridization events due to climate and environmental changes).

Dichotomous Keys Based on Cytogenetic Data for Triatomines Reported in Brazilian Regions with Outbreaks of Orally Transmitted Chagas Disease (Pernambuco and Rio Grande Do Norte).

Chagas disease (CD) affects about eight million people worldwide. Brazil has the highest number of estimated cases and the largest number of deaths due to CD. Considering the recent outbreaks of oral CD involving at least 27 cases of acute CD in Pernambuco (PE) as well as 18 cases and 2 deaths in the Rio Grande do Norte (RN), we developed dichotomous keys for the identification of triatomine species in these Brazilian states based on cytogenetic data. All triatomine species could be distinguished by cytogenetic characteristics, emphasizing the importance of the newly developed taxonomic keys for the correct identification of triatomes from PE and RN, particularly for species that exhibit morphological similarities, such as Triatoma brasilensis and T. petrocchiae (present in both states) and T. maculata and T. pseudomaculata (as T. pseudomaculata has been misidentified as T. maculata in PE and RN). These alternative keys are expected to provide a useful tool for the scientific community and, above all, health agents, aimed at preventing mistakes from occurring in the identification of the vectors present in PE and RN related to CD outbreaks caused by oral infection.

Triatominae (Hemiptera: Reduviidae) fauna in bird nests: the case of Morrinhos, Ceará, Brazil.

Triatominae are associated with various Brazilian habitats, including bird nests, animal burrows, and peridomestic structures. Despite extensive studies on triatomines in domiciliary environments in Ceará, Brazil, there has been limited research on their presence in the wild. This study focuses on the municipality of Morrinhos in Ceará, which is characterized by a Caatinga biome and riparian forests along the Acaraú River. During the study, a total of 185 nests of Pseudoseisura cristata were analyzed in rural locations and triatomines were collected in 26 nests from 12 different locations. A total of 117 triatomines was collected, belonging to three species: Psammolestes tertius, Rhodnius nasutus, and Triatoma pseudomaculata. Rhodnius nasutus was the only species found in a nest inhabited by Didelphis albiventris. Nests of P. cristata serve as shelters for various animals, providing an ideal environment for triatomines to establish colonies due to their proximity to food sources and their generalist feeding habits. The incorporation of anthropogenic materials by birds in nest construction can indirectly affect the presence of other animals that may serve as food sources for triatomines. Understanding the interactions between triatomines, birds, and their habitats is essential for assessing the risks associated with triatomine infestation in wild ecosystems.

Exploring the Hidden World of Vectors of Chagas Disease: A Fascinating Look at the Taxonomic Aspects of the Psammolestes Genus (Hemiptera, Triatominae).

Chagas disease (CD) is a neglected illness affecting approximately seven million individuals, with vector transmission occurring via triatomine bugs. The Rhodniini tribe comprises 24 species, grouped into the Rhodnius and Psammolestes genera. Given the importance of accurately identifying CD vectors, the taxonomy of Psammolestes spp. was revisited using morphological and morphometric data. Specimens of P. tertius, P. coreodes, and P. arthuri were collected, and the morphological characteristics of the head, thorax, abdomen, and eggs were analyzed. Morphometric studies of eggs were also conducted. Dichotomous keys allowing for the differentiation of Psammolestes spp. were elaborated based on adult insect and egg morphological characteristics. Through these studies, it was possible to differentiate the three Psammolestes species and confirm that this genus should not be classified under the Rhodnius genus, contributing to Rhodniini taxonomy.

Cytogenetic Key to Identification of Triatominae Species Reported in an Outbreak Region of Oral Transmission of Chagas Disease in the Brazilian Northeast.

Oral transmission from the consumption of processed food with triatomines and/or their feces infected with Trypanosoma cruzi prevails among recent cases of Chagas disease in Brazil. In Paraíba, a state of the Brazilian northeast, there was an outbreak caused by the consumption of sugarcane juice that resulted in 26 cases of infection and one death. Until now, 10 species of triatomines have been reported in this Brazilian state. Thus, we developed a dichotomous key to assist in the correct identification of Paraíba triatomines based on cytogenetic data. The dichotomous key allowed the differentiation of all the species in this state. Although the purpose of CytoKeys is not to replace dichotomous keys based on morphological data, the use of these complementary keys can help to solve taxonomic problems, preventing identification errors, especially between similar species such as Triatoma brasiliensis and Triatoma petrocchiae, both present in the Brazilian northeast.

First evidence of gonadal hybrid dysgenesis in Chagas disease vectors (Hemiptera, Triatominae): gonad atrophy prevents events of interspecific gene flow and introgression.

BACKGROUND: Hybridization events between Triatoma spp. have been observed under both natural and laboratory conditions. The ability to produce hybrids can influence different aspects of the parent species, and may even result in events of introgression, speciation and extinction. Hybrid sterility is caused by unviable gametes (due to errors in chromosomal pairing [meiosis]) or by gonadal dysgenesis (GD). All of the triatomine hybrids analyzed so far have not presented GD. We describe here for the first time GD events in triatomine hybrids and highlight these taxonomic and evolutionary implications of these events. METHODS: Reciprocal experimental crosses were performed between Triatoma longipennis and Triatoma mopan. Intercrosses were also performed between the hybrids, and backcrosses were performed between the hybrids and the parent species. In addition, morphological and cytological analyzes were performed on the atrophied gonads of the hybrids. RESULTS: Hybrids were obtained only for the crosses T. mopan♀ × T. longipennis♂. Intercrosses and backcrosses did not result in offspring. Morphological analyses of the male gonads of the hybrids confirmed that the phenomenon that resulted in sterility of the hybrid was bilateral GD (the gonads of the hybrids were completely atrophied). Cytological analyses of the testes of the hybrids also confirmed GD, with no germ cells observed (only somatic cells, which make up the peritoneal sheath). CONCLUSIONS: The observations made during this study allowed us to characterize, for the first time, GD in triatomines and demonstrated that gametogenesis does not occur in atrophied gonads. The characterization of GD in male hybrids resulting from the crossing of T. mopan♀ × T. longipennis♂ highlights the importance of evaluating both the morphology and the cytology of the gonads to confirm which event resulted in the sterility of the hybrid: GD (which results in no gamete production) or meiotic errors (which results in non-viable gametes).

Climate and Environmental Changes and Their Potential Effects on the Dynamics of Chagas Disease: Hybridization in Rhodniini (Hemiptera, Triatominae).

Chagas disease affects about eight million people. In view of the issues related to the influence of anthropogenic changes in the dynamics of the distribution and reproductive interaction of triatomines, we performed experimental crosses between species of the Rhodniini tribe in order to evaluate interspecific reproductive interactions and hybrid production capacity. Reciprocal crossing experiments were conducted among Rhodnius brethesi × R. pictipes, R. colombiensis × R. ecuadoriensis, R. neivai × R. prolixus, R. robustus × R. prolixus, R. montenegrensis × R. marabaensis; R. montenegrensis × R. robustus, R. prolixus × R. nasutus and R. neglectus × R. milesi. With the exception of crosses between R. pictipes ♀ × R. brethesi ♂, R. ecuadoriensis ♀ × R. colombiensis ♂ and R. prolixus ♀ × R. neivai ♂, all experimental crosses resulted in hybrids. Our results demonstrate that both allopatric and sympatric species produce hybrids, which can generate concern for public health agencies in the face of current anthropogenic events. Thus, we demonstrate that species of the Rhodniini tribe are capable of producing hybrids under laboratory conditions. These results are of great epidemiological importance and raise an important discussion about the influence of climatic and environmental interactions on Chagas disease dynamics.

Transcriptomics Applied to the Study of Chagas Disease Vectors.

Chagas disease is a neglected disease caused by the protozoan Trypanosoma cruzi, and is transmitted mainly by the feces of contaminated triatomines. Knowledge of the biological, ecological, behavioral, genetic, taxonomic, and systematic aspects of these vectors can contribute to the planning of vector control programs, because all species are considered to be potential vectors of Chagas disease. Transcriptomic studies, in general, provided a new view of the physiology of triatomines (aiding in the knowledge of reproductive aspects of the hematophagy process and even the immune system and the sensory apparatus) and even contributed, as a new tool, to the taxonomy and systematics of these insects. Thus, we conducted a review of the transcriptomic studies on Chagas disease vectors.

Phylogenetic relationships and evolutionary patterns of the genus Psammolestes Bergroth, 1911 (Hemiptera: Reduviidae: Triatominae).

BACKGROUND: The evolutionary history of biodiversity in South America has been poorly studied in the seasonal dry tropical forest (SDTF). Species diversification in this ecosystem may have a twofold explanation. First, intermittent connections in the middle and late Pleistocene promoted species dispersal and/or genetic connectivity between lineages isolated in disjunct patches of forest. Second, allopatric speciation proceeded immediately after the formation and colonization of the SDTF in the Neogene. Here we studied the diversification of Psammolestes, a genus endemic of the SDTF and naturally infected with Trypanosoma cruzi (agent of Chagas disease), using a combination of phylogenetic, population genetics and niche model methods, and evaluated the reliability of the three morphospecies currently recognized. RESULTS: Our multilocus analyses recovered P. coreodes and P. tertius in a monophyletic clade sister to P. arthuri. Species delimitation tests recovered these lineages as different species despite the shared genetic variation observed between P. coreodes and P. tertius in five genes. Also, genetic variation of the genus clustered in three groups that were consistent with the three morphospecies. Our demographic model predicted a scenario of divergence in absence of gene flow, suggesting that mixed haplotypes may be the result of shared ancestral variation since the divergence of the subtropical-temperate species P. coreodes and P. tertius. In contrast, the tropical species P. arthuri was highly differentiated from the other two in all tests of genetic structure, and consistently, the Monmonier's algorithm identified a clear geographical barrier that separates this species from P. coreodes and P. tertius. CONCLUSIONS: We found three genetically structured lineages within Psammolestes that diverged in absence of gene flow in the late Miocene. This result supports a scenario of species formation driven by geographical isolation rather than by divergence in the face of gene flow associated with climatic oscillations in the Pleistocene. Also, we identified the Amazon basin as a climatic barrier that separates tropical from subtropical-temperate species, thus promoting allopatric speciation after long range dispersion. Finally, each species of Psammolestes occupies different climatic niches suggesting that niche conservatism is not crucial for species differentiation. These findings influence the current vector surveillance programs of Chagas disease in the region.

Trends in evolution of the Triatomini tribe (Hemiptera, Triatominae): reproductive incompatibility between four species of geniculatus clade.

BACKGROUND: The geniculatus clade, composed by the rufotuberculatus, lignarius, geniculatus and megistus groups, relates evolutionarily the species of the genus Panstrongylus and Nesotriatoma. Several studies have shown that triatomine hybrids can play an important role in the transmission of Chagas disease. Natural hybrids between species of the geniculatus clade have never been reported to our knowledge. Thus, carrying out experimental crosses between species of the geniculatus clade can help to elucidate the taxonomic issues as well as contribute to the epidemiological knowledge of this group. METHODS: Experimental crosses were carried out between species of the megistus and lignarius groups to evaluate the reproductive compatibility between them. A phylogenetic reconstruction was also performed with data available in GenBank for the species of the geniculatus clade to show the relationships among the crossed species. RESULTS: Phylogenetic analysis grouped the species of the geniculatus clade into four groups, as previously reported. In the interspecific crosses performed there was no hatching of eggs, demonstrating the presence of prezygotic barriers between the crossed species and confirming their specific status. CONCLUSIONS: In contrast to the other groups of the Triatomini tribe, as well as the Rhodniini, there are prezygotic barriers that prevent the formation of hybrids between species of the megistus and lignarius groups. Thus, the geniculatus clade may represent an important evolutionary model for Triatominae, highlighting the need for further studies with greater sample efforts for this clade (grouping the 17 species of Panstrongylus and the three of Nesotriatoma).

Do not judge a book by its cover: would Triatoma tibiamaculata (Pinto, 1926) belong to Triatoma Laporte, 1832, or to Panstrongylus Berg, 1879, with misleading homoplasies?

BACKGROUND: Triatoma tibiamaculata is a species distributed in ten Brazilian states which has epidemiological importance as it has already been found infecting household areas. The taxonomy of this triatomine has been quite unstable: it was initially described as Eutriatoma tibiamaculata. Later, the species was transferred from the genus Eutriatoma to Triatoma. Although included in the genus Triatoma, the phylogenetic position of T. tibiamaculata in relation to other species of this genus has always been uncertain once this triatomine was grouped in all phylogenies with the genus Panstrongylus, rescuing T. tibiamaculata and P. megistus as sister species. Thus, we evaluated the generic status of T. tibiamaculata using phylogenetic and chromosomal analysis. METHODS: Chromosomal (karyotype) and phylogenetic (with mitochondrial and nuclear markers) analyses were performed to assess the relationship between T. tibiamaculata and Panstrongylus spp. RESULTS: The chromosomal and phylogenetic relationship of T. tibiamaculata and Panstrongylus spp. confirms the transfer of the species to Panstrongylus with the new combination: Panstrongylus tibiamaculatus. CONCLUSIONS: Based on chromosomal and phylogenetic characteristics, we state that P. tibiamaculatus comb. nov. belongs to the genus Panstrongylus and that the morphological features shared with Triatoma spp. represent homoplasies.

Morphological, Cytological and Molecular Studies and Feeding and Defecation Pattern of Hybrids from Experimental Crosses between Triatoma sordida and T. rosai (Hemiptera, Triatominae).

Under laboratory conditions, Triatoma rosai and T. sordida are able to cross and produce hybrids. In the face of climate and environmental changes, the study of hybrids of triatomines has evolutionary and epidemiological implications. Therefore, we performed morphological, cytological and molecular studies and characterized the feeding and defecation pattern of hybrids from crosses between T. sordida and T. rosai. The morphological characterization of the female genitalia of the hybrids showed that characteristics of both parental species segregated in the hybrids. Cytogenetic analyzes of hybrids showed regular metaphases. According to molecular studies, the mitochondrial marker Cytochrome B (CytB) related the hybrids with T. sordida and the nuclear marker Internal Transcribed Spacer 1 (ITS-1) related the hybrids with T. rosai. Both parents and hybrids defecated during the blood meal. Thus, the hybrids resulting from the cross between T. sordida and T. rosai presented segregation of phenotypic characters of both parental species, 100% homeology between homeologous chromosomes, phylogenetic relationship with T sordida and with T. rosai (with CytB and ITS-1, respectively), and, finally, feeding and defecation patterns similar to the parents.

Chagas Disease Vectors of Espírito Santo, Brazil: First Report of Triatoma infestans (Klug, 1834) (Hemiptera, Triatominae) in the Brazilian State and Development of an Identification Key Based on Cytogenetic Data.

Chagas disease is a potentially life-threatening illness caused by the protozoan Trypanosoma cruzi and transmitted, mainly, by hematophagous insects of the Triatominae subfamily. In Brazil, there are currently about 66 triatomine species distributed throughout the country's 27 states. Triatoma infestans is considered as a species of great vectorial importance, mainly because of its biological characteristics, such as the high degree of anthropophilia, adaptation to the home environment, ability to withstand long periods of fasting, and present a wide geographical distribution. Taking into account the epidemiological importance of these species, we carried out the first report of T. infestans in the Espírito Santo, Brazil, and development of an identification key for all species notified in that state, based on cytogenetic data. This information is important because they contribute to the direction of epidemiological surveillance activities carried out by vector control programs of the Espírito Santo, Brazil.

Trends in Taxonomy of Chagas Disease Vectors (Hemiptera, Reduviidae, Triatominae): From Linnaean to Integrative Taxonomy.

Chagas disease is a neglected tropical disease caused by the protozoan Trypanosoma cruzi and transmitted mainly by members of the subfamily Triatominae. There are currently 157 species, grouped into 18 genera and five tribes. Most descriptions of triatomine species are based on classical taxonomy. Facing evolutionary (cryptic speciation and phenotypic plasticity) and taxonomic (more than 190 synonymizations) problems, it is evident that integrative taxonomy studies are an important and necessary trend for this group of vectors. Almost two-and-a-half centuries after the description of the first species, we present for the first time the state-of-the-art taxonomy of the whole subfamily, covering from the initial classic studies to the use of integrative taxonomy.

Biological, ecological, morphological and cytogenetic analyses, with taxonomic notes of emZelurus/em emochripennis/em (Stål, 1854) (Hemiptera: Heteroptera: Reduviidae: Reduviinae).

Based on specimens of Zelurus ochripennis (Stål, 1854) collected in the entrance zone of a small terrestrial cave, some biological, ecological, morphological and cytogenetic data, as well, as the general morphology of their nymphs are presented for the first time. A short taxonomic summary of the species is also provided.