The use of subtle features as species diagnostic traits in taxa with high morphological similarity sometimes fails in discriminating intraspecific variation from interspecific differences, leading to an incorrect species delimitation. A clear assessment of species boundaries is particularly relevant in disease vector organisms in order to understand epidemiological and evolutionary processes that affect transmission capacity. Here, we assess the validity of the recently described Rhodnius taquarussuensis (da Rosa et al., 2017) using interspecific crosses and molecular markers. We did not detect differences in hatching rates in interspecific crosses between R. taquarussuensis and R. neglectus (Lent, 1954). Furthermore, genetic divergence and species delimitation analyses show that R. taquarussuensis is not an independent lineage in the R. prolixus group. These results suggest that R. taquarussuensis is a phenotypic form of R. neglectus instead of a distinct species. We would like to stress that different sources of evidence are needed to correctly delimit species. We consider this is an important step in understanding vectorial Chagas disease spread and transmission.

Chagas disease is the most prevalent neglected tropical disease in the Americas and makes an important contribution to morbidity and mortality rates in countries where it is endemic since 30 to 40% of patients develop cardiac diseases, gastrointestinal disorders, or both. In this paper, a new species of the genus Triatoma is described based on specimens collected in the Department San Miguel, Province of Corrientes, Argentina. Triatoma rosai sp. nov. is closely related to T. sordida (Stål, 1859), and was characterized based on integrative taxonomy using morphological, morphometric, molecular data, and experimental crosses. These analyses, combined with data from the literature (cytogenetics, electrophoresis pattern, molecular analyses, cuticular hydrocarbons pattern, geometric morphometry, cycle, and average time of life as well as geographic distribution) confirm the specific status of T. rosai sp. nov. Natural Trypanosoma cruzi infection, coupled with its presence mostly in peridomestic habitats, indicates that this species can be considered as an important Chagas disease vector from Argentina.

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.

Background The tribe Rhodniini is a monophyletic group composed of 24 species grouped into two genera: Rhodnius and Psammolestes. The genus Psammolestes includes only three species, namely P. coreodes, P. tertius and P. arthuri. Natural hybridization events have been reported for the Rhodniini tribe (for genus Rhodnius specifically). Information obtained from hybridization studies can improve our understanding of the taxonomy and systematics of species. Here we report the results from experimental crosses performed between P. tertius and P. coreodes and from subsequent analyses of the reproductive and morphological aspects of the hybrids. Methods Crossing experiments were conducted between P. tertius and P. coreodes to evaluate the pre- and post-zygotic barriers between species of the Rhodniini tribe. We also performed cytogenetic analyses of the F1 hybrids, with a focus on the degree of pairing between the homeologous chromosomes, and morphology studies of the male gonads to evaluate the presence of gonadal dysgenesis. Lastly, we analyzed the segregation of phenotypic characteristics. Results Interspecific experimental crosses demonstrated intrageneric genomic compatibility since hybrids were produced in both directions. However, these hybrids showed a high mortality rate, suggesting a post-zygotic barrier resulting in hybrid unviability. The F1 hybrids that reached adulthood presented the dominant phenotypic segregation pattern for P. tertius in both directions. These insects were then intercrossed; the hybrids were used in the cross between P. tertius ♀ × P. coreodes ♂ died before oviposition, and the F1 hybrids of P. coreodes ♀ x P. tertius ♂ oviposited and their F2 hybrids hatched (however, all specimens died after hatching, still in first-generation nymph stage, pointing to a hybrid collapse event). Morphological analyses of male gonads from F1 hybrids showed that they did not have gonadal dysgenesis. Cytogenetic analyses of these triatomines showed that there were metaphases with 100% pairing between homeologous chromosomes and metaphases with pairing errors. Conclusion The results of this study demonstrate that Psammolestes spp. have intrageneric genomic compatibility and that post-zygotic barriers, namely unviability of hybrid and hybrid collapse, resulted in the breakdown of the hybrids of P. tertius and P. coreodes, confirming the specific status of species based on the biological concept of species. Graphical abstract

ABSTRACT. The Triatominae subfamily comprises 6 tribes. The tribe Rhodniini comprises 2 genera and 22 nominal species. Rhodnius montenegrensis (Hemiptera, Triatominae) was recently described as evolutionarily related to R. robustus. Therefore, in order to contribute to karyosystematic study of the tribe Rhodniini, this report describes the number of chromosomes and compares the karyotype of R. montenegrensis to that of all other species in the tribe, in order to determine the karyotypic evolution of the tribe Rhodniini. The seminiferous tubules of adult males, after being removed and fixated on a cover slip, were processed with lactoaceto-orcein for cytogenetic analysis. R. montenegrensis, as well as all other species of the tribe Rhodniini showed 22 chromosomes (20 autosomes + XY). Thus, we hereby describe the karyotype of the species R. montenegrensis and mainly highlight that the Karyotype of Rhodnius montenegrensis tribe Rhodniini displays karyotypic homogeneity, demonstrating itself as a derived group to a lesser extent when compared to the number of chromosomes of the common ancestors of the subfamily Triatominae.