High chromosomal mobility of rDNA clusters in holocentric chromosomes of Triatominae, vectors of Chagas disease (Hemiptera-Reduviidae).

The subfamily Triatominae (Hemiptera-Reduviidae) includes more than 150 blood-sucking species, potential vectors of the protozoan Trypanosoma cruzi, causative agent of Chagas disease. A distinctive cytogenetic characteristic of this group is the presence of extremely stable chromosome numbers. Unexpectedly, the analyses of the chromosomal location of ribosomal gene clusters and other repetitive sequences place Triatominae as a significantly diverse hemipteran subfamily. Here, we advance the understanding of Triatominae chromosomal evolution through the analysis of the 45S rDNA cluster chromosomal location in 92 Triatominae species. We found the 45S rDNA clusters in one to four loci per haploid genome with different chromosomal patterns: On one or two autosomes, on one, two or three sex chromosomes, on the X chromosome plus one to three autosomes. The movement of 45S rDNA clusters is discussed in an evolutionary context. Our results illustrate that rDNA mobility has been relatively common in the past and in recent evolutionary history of the group. The high frequency of rDNA patterns involving autosomes and sex chromosomes among closely related species could affect genetic recombination and the viability of hybrid populations, which suggests that the mobility of rDNA clusters could be a driver of species diversification.

Reducing interruptions during duty radiology shifts, assessment of its benefits and review of factors affecting the radiology working environment.

AIM: To investigate and reduce the number of inappropriate interruptions to the duty radiology registrar, as well as subjectively assess the effect of reducing interruptions and identify other methods of improving the duty working environment. MATERIALS AND METHODS: A pre- and post-intervention prospective quantitative study and post-intervention retrospective qualitative study of duty radiology registrars was performed at a tertiary referral centre. The first cycle of the quantitative study was performed prior to implementation of a telephone triage system. The second cycle and qualitative study were performed afterwards. RESULTS: The average number of interruptions per day dropped by 43.7% after the intervention. Moreover, inappropriate interruptions dropped from one in three to one in five interruptions. Improvement was demonstrated following the intervention for perceived patient safety, workload, reporting efficiency, reporting accuracy, work satisfaction, and stress. The most common hindrance to a good working environment was interruptions (36%). The most common suggestion for improvement was improved comfort (33%). CONCLUSION: The present study demonstrates that a telephone triage system can substantially reduce the number of interruptions to the duty radiologist. It also demonstrates that reducing interruptions in radiology has the potential to improve the working environment in many ways.

New chromosomal evidence for the origin of Triatoma infestans populations from Brazil.

In this study, the karyometry of different Triatoma infestans populations from different states of Brazil was analyzed and compared with those of a population from Cochabamba. No significant differences were found between the population from Cochabamba and those from Brazil. These results are consistent with the origin of the T. infestans populations of Brazil by a founder effect from Cochabamba. Moreover, these findings also confirm that a founder effect occurred during the dispersal of T. infestans populations in different Brazilian states.

New record and cytogenetic analysis of Psammolestes tertius Lent & Jurberg, 1965 (Hemiptera, Reduviidae, Triatominae) from Bahia State, Brazil.

This paper reports on the first occurrence of Psammolestes tertius in the Chapada Diamantina region, located in the city of Seabra, Bahia State, in northeastern Brazil. Following an active search, 24 P. tertius specimens were collected from Phacellodomus rufifrons (rufous-fronted thornbird) nests. The insects did not present any symptoms of infection by Trypanosoma cruzi. P. tertius males were cytogenetically analyzed, and the results were compared with those of other specimens from the Brazilian State of Ceará. Triatomines from both locations presented the same cytogenetic characteristics: 22 chromosomes, little variation in the size of the autosomes, Y chromosomes that were larger than the X chromosomes, a chromocenter formed only by the sex chromosomes during prophase, and autosomes lacking constitutive heterochromatin. However, it is important to note that this species shows intraspecific chromosomal variation. In light of the results obtained, it is recommended that more studies be performed to characterize P. tertius. These studies will be particularly helpful in understanding this species in ecological, biological, biogeographical, and phylogenetic terms.

Presence of chromatoid bodies in the Rhodnius genus detected by cytochemical analysis.

In addition to the epidemiological importance regarding the transmission of Chagas disease, triatomines are also important biological models for cellular studies, because they have holocentric chromosomes, post-reductional meiosis for sex chromosomes, and nucleolar persistence. Although the nucleolus is present during spermiogenesis, it was suggested that it is inactivated and all transcriptional activity necessary for cell differentiation is supported by the chromatoid body (CB). Thus, considering the importance of CB to triatomine spermatogenesis, this paper aims to analyze the presence of this organelle in the Rhodnius genus. The testicles of five adult R. neglectus and R. prolixus males were analyzed after semi-fine sections were performed, and the material was subjected to silver ion impregnation. The CB organelle was found in the spermatid of both species. Most CB studies focused on the Triatoma genus. This study described the existence of CB in Rhodnius, confirming thus the importance of this cytoplasmic organelle to spermiogenesis of these insect vectors of Chagas disease.

Spermiotaxonomy of the tribe Rhodniini (Hemiptera, Triatominae).

The tribe Rhodniini is a monophyletic group composed of 22 species, with 19 in the Rhodnius genus and three in the Psammolestes genus. These insects are morphologically very similar (cryptic species), and new tools are important for investigating the taxonomy of these vectors. Spermiotaxonomy is an important tool in differentiating between related species, and this study analyzed the spermatids of Rhodniini species to elucidate their spermiotaxonomy. All of the Rhodniini species contained two heteropyknotic filaments in the extremities of their cells. Although spermiotaxonomy has been an important tool in differentiating between species of the Triatoma genus, all of the species in the Rhodnius genus exhibited the same characteristics in their male gametes. However, spermatid analysis made it possible to confirm the monophyly of the Rhodniini tribe, because Psammolestes tertius had the same pattern as that described for Rhodnius. The results of this study demonstrate that spermiotaxonomy, in addition to being an important tool for differentiating between related species of Triatoma, can be used as an optimization tool in phylogenetic analyses.

Insights into a hotspot in the Brasiliensis subcomplex (Hemiptera, Triatominae) by analysis of D2 domain of the nuclear gene 28S.

The Brasiliensis subcomplex is a monophyletic group formed by the species Triatoma brasiliensis brasiliensis, T. b. macromelasoma, T. juazeirensis, T. melanica, and T. sherlocki. However, using cytogenetic data and experimental hybrid crosses, T. lenti and T. petrochiae were also grouped into this subcomplex. This study aims to analyze the properties of hotspot in the D2 domain of the nuclear gene 28S in all species of the Brasiliensis subcomplex as well as T. lenti and T. petrochiae. These species show two transversions at position 385 (G↔C and T↔G). We suggest that this mutation in haplotype 4 may be an initial molecular tool that supports the relationship of these species with the subcomplex. In addition to the transversion at haplotype 4, these species, aside from T. melanica, also possess a transversion at position 385 (G↔T) in haplotype 1. Thus, we describe the hotspot mutations of the D2 domain of the nuclear gene 28S for species in Brasiliensis subcomplex as follows: three transversions are present at position 385 of haplotypes 1 and 4, which are shared by members of the subcomplex as well as T. lenti and T. petrochiae. These transversions may be considered a synapomorphy between these species. However, we emphasize that new phylogenetic studies should be conducted to evaluate whether T. lenti and T. petrochiae are truly members of the Brasiliensis subcomplex.

Nucleolar-persistence phenomenon during spermatogenesis in genus Meccus (Hemiptera, Triatominae).

The Triatominae subfamily consists of 150 species in 18 genera, grouped into six tribes. In cytogenetics, triatomines are important biological models because they have holocentric chromosomes and nucleolar persistence in meiosis. The phenomenon of nucleolar persistence has been described for 23 species of triatomine in three genera: Triatoma, Rhodnius, and Panstrongylus. However, new species and genera should be analyzed to assess whether nucleolar persistence is a peculiarity of Triatominae. Thus, this study aimed to analyze nucleolar behavior during spermatogenesis of Meccus pallidipennis and M. longipennis, focusing on the nucleolar-persistence phenomenon. Through the analysis of spermatogenesis, more specifically of meiotic metaphase, we observed the phenomenon of nucleolar persistence in M. pallidipennis and M. longipennis, represented by remnants of nucleolar material in metaphase. Thus, although nucleologenesis of new species, and, especially, new genera, should be analyzed, this study confirms for the first time the phenomenon of nucleolar persistence in the genus Meccus. Therefore, we emphasize the importance of new studies in this area in order to assess whether this phenomenon is truly a synapomorphy of these hematophagous insects.

Cytogenetic analysis of Triatoma pseudomaculata Corrêa and Espínola, 1964 (Hemiptera, Triatominae) from different Brazilian states.

Triatoma maculata and T. pseudomaculata are allopatric; however, it is believed that T. maculata was introduced into the Brazilian northeast by passive transportation of the nymphs between the feathers of migratory birds, followed by the speciation of T. pseudomaculata. T. pseudomaculata is the second most epidemiologically important species in the northeast of Brazil after T. brasiliensis. Therefore, given the broad range of T. pseudomaculata, the objective of the present study was to cytogenetically analyze different populations of T. pseudomaculata from different Brazilian states in order to investigate possible intraspecific chromosomal variation between them. Three adult T. pseudomaculata males from each population (Pernambuco, Ceará, Paraíba, Bahia, Rio Grande do Norte, and Piauí) were analyzed by lacto-acetic orcein and C-banding. All of the specimens analyzed exhibited the same cytogenetic characteristics, i.e., 22 chromosomes (20 autosomes and XY), a chromocenter formed by the X and Y sex chromosomes and one pair of autosomes, and heterochromatin blocks in three or four pairs of autosomes. These data confirm that all of the populations analyzed were T. pseudomaculata, and although they may be subject to different selection pressures they have maintained the genetic integrity that characterizes the species.

Comparing de novo and reference-based transcriptome assembly strategies by applying them to the blood-sucking bug Rhodnius prolixus.

High Throughput Sequencing capabilities have made the process of assembling a transcriptome easier, whether or not there is a reference genome. But the quality of a transcriptome assembly must be good enough to capture the most comprehensive catalog of transcripts and their variations, and to carry out further experiments on transcriptomics. There is currently no consensus on which of the many sequencing technologies and assembly tools are the most effective. Many non-model organisms lack a reference genome to guide the transcriptome assembly. One question, therefore, is whether or not a reference-based genome assembly gives better results than de novo assembly. The blood-sucking insect Rhodnius prolixus-a vector for Chagas disease-has a reference genome. It is therefore a good model on which to compare reference-based and de novo transcriptome assemblies. In this study, we compared de novo and reference-based genome assembly strategies using three datasets (454, Illumina, 454 combined with Illumina) and various assembly software. We developed criteria to compare the resulting assemblies: the size distribution and number of transcripts, the proportion of potentially chimeric transcripts, how complete the assembly was (completeness evaluated both through CEGMA software and R. prolixus proteome fraction retrieved). Moreover, we looked for the presence of two chemosensory gene families (Odorant-Binding Proteins and Chemosensory Proteins) to validate the assembly quality. The reference-based assemblies after genome annotation were clearly better than those generated using de novo strategies alone. Reference-based strategies revealed new transcripts, including new isoforms unpredicted by automatic genome annotation. However, a combination of both de novo and reference-based strategies gave the best result, and allowed us to assemble fragmented transcripts.

First cytogenetic study of Cavernicola pilosa Barber, 1937 (Hemiptera, Triatominae).

Cavernicola pilosa is a triatomine species that lives in caves and feeds on bat blood. This vector has a wide geographical distribution, and is found in Brazil, Colombia, Panama, Peru, and Venezuela. Little is known about the reproductive biology of this species, because most previous studies have only characterized its morphology, morphometry, ecology, and epidemiology. Therefore, this study aimed to obtain preliminary data related to spermatogenesis in C. pilosa by conducting cytogenetic analysis. Analysis of the heterochromatic pattern of C. pilosa during the initial prophases revealed that heterochromatic blocks are only present in the sex chromosomes. Based on the analyses of the meiotic metaphase and prophases, we found that the sex determination system of C. pilosa is XY and the chromosomes are holocentric. C. pilosa spermatids are filamentous and have long flagella. It was not possible to detect corpuscle or filament heteropycnosis in spermatids of this species. The initial cytogenetic data presented in this study are important in characterizing the spermatogenesis and heterochromatic patterns of C. pilosa. Our results suggest that adaptation to troglodytism did not result in differences in spermatogenesis in this vector.

Chromosomal evolution in the pallescens group (Hemiptera, Triatominae).

Rhodnius colombiensis, in conjunction with R. pallescens and R. ecuadoriensis, forms the monophyletic pallescens group. Cytogenetic analyses of these closely related species would further our understanding of the taxonomy and evolution of this group. In this study, R. colombiensis was cytogenetically analyzed, and the results were compared with cytogenetic data from other species of the pallescens group, particularly their chromosomal evolution. We found that this triatomine has heteropycnotic blocks in five autosomal bivalents at both metaphase I and II. The derivation of R. colombiensis from R. pallescens led to significant loss of heteropycnotic and heterochromatic regions (approximately 50%). R. ecuadoriensis is the most differentiated of the group because it has lost all heterochromatin and heteropyknotic blocks in the autosomes. Based on the heteropyknotic and heterochromatic pattern of R. colombiensis and the chromosomal evolution analysis of the pallescens group, we suggest that the karyotype of R. colombiensis and R. ecuadoriensis lost its heteropycnotic and heterochromatic blocks during speciation. Furthermore, this loss could be related to adaptation to different environments.

Karyotype of Rhodnius montenegrensis (Hemiptera, Triatominae).

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 lacto-aceto-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 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.

Diploid chromosome set of kissing bug Triatoma baratai (Hemiptera, Triatominae).

Triatomines are insects that are taxonomically included in the Hemiptera order and Triatominae subfamily. Based on phenotypic similarity, capacity hybridization, and genetic and ecological aspects, the triatomine species can be grouped into specific complexes and subcomplexes. However, these groupings have not been confirmed. Cytogenetic analyses are important cytotaxonomic tools for improving the taxonomic knowledge of triatomines. Thus, we examined the karyotype of Triatoma baratai and compared the results with those of other species in the Matogrossensis subcomplex in order increase the understanding of vector potential. We also examined the cytotaxonomic classification of this insect. Triatoma baratai, similarly to other species that currently compose the Matogrossensis subcomplex, contains 22 chromosomes (20A + XY). Here, we describe the diploid chromosome set of T. baratai. We confirmed their current classification in the Matogrossensis subcomplex and demonstrated that the species in this subcomplex present karyotype homogeneity.

Distribution of constitutive heterochromatin in species of triatomines with fragmentation of sex chromosomes X.

Cytogenetic analyses of triatomines are considered to be important taxonomic tools. Thus, we analyzed the pattern of constitutive heterochromatin in 7 species of triatomine with fragmentation of the sex chromosome X, focusing on the cytotaxonomy of these triatomines. The species analyzed included Triatoma vitticeps, Triatoma melanocephala, Triatoma tibiamaculata, Triatoma protracta, Meccus pallidipennis, Panstrongylus megistus, and Panstrongylus lignarius. The seminiferous tubules of the adult males were subjected to C-banding. P. megistus and P. lignarius showed differences in chromosome number and disposition of constitutive heterochromatin, as only P. lignarius showed C-blocks in autosomes. C-banding can differentiate these species, since one of the sex chromosome (X) is heterochromatic in T. vitticeps. T. protracta showed C-blocks in both ends of all autosomes, T. tibiamaculata showed terminal C-dots in some autosomal pairs and M. pallidipennis did not show constitutive heterochromatin in autosomes. Thus, we confirmed the heterochromatic pattern of 7 species of insects and emphasized the importance of cytogenetic techniques for C-banding for taxonomy studies of the triatomines, which are important vectors of Chagas disease.

Distribution of constitutive heterochromatin in Triatoma melanocephala (Hemiptera, Triatominae).

In principle, Triatoma melanocephala was included in the Brasiliensis subcomplex on the basis of morphological parameters and geographical layout, since there were no other relevant data available in the literature. On the basis of karyotype, it has been proposed to exclude T. melanocephala, as well as of T. vitticeps and T. tibiamaculata, from the subcomplex, which shows fragmentation of the X sex chromosomes, thereby approaching the species of North America. Therefore, the present study aimed to determine the pattern of constitutive heterochromatin of T. melanocephala to provide new data on the cytotaxonomy of this vector of Chagas disease and especially to try to relate this species to some group, complex or subcomplex of triatomine species, aiding in their classification. This species showed no constitutive heterochromatin in the autosomes or X sex chromosome, but only the Y sex chromosome. The number of chromosomes and heterochromatin pattern of T. melanocephala proved to be identical to that described for Panstrongylus lutzi. Thus, the present study demonstrated a tentative relationship between T. melanocephala and P. lutzi. However, we emphasize that other comparative studies should be conducted between these species, such as experimental crosses and molecular, enzymatic, morphological, and morphometric analyses to determine whether these species are actually evolutionarily related or if the number of chromosomes and the heterochromatin pattern emerged as homoplasies in T. melanocephala and P. lutzi.

Cross-species transferability of eastern white pine (Pinus strobus) nuclear microsatellite markers to five Mexican white pines.

We examined cross-species transferability and usefulness of six nuclear microsatellite markers developed in consubgeneric eastern white pine (Pinus strobus) with regard to ecologically and commercially important Mexican white pine species of conservation genetics concern: Pinus chiapensis (Mart.) Andresen, P. flexilis James, P. strobiformis Engelm., P. ayacahuite Ehrenb. Ex Schltdl, and P. ayacahuite var. veitchii (Roezl) G.R. Shaw. Four to six microsatellite loci were found to be polymorphic in different species, with moderate to high informativeness in a relatively small number of samples (PIC/HE=0.25-0.93). This successful transfer sidesteps the time- and resource-consuming development of species-specific microsatellite markers, and will facilitate population and conservation genetic studies and genetic resource management of the less studied Mexican white pines.