Revalidation and redescription of Triatoma brasiliensis macromelasoma Galv?o, 1956 and an identification key for the Triatoma brasiliensis complex (Hemiptera: Reduviidae: Triatominae)

Triatoma brasiliensis macromelasoma is revalidated based on the results of previous multidisciplinary studies on the Triatoma brasiliensis complex, consisting of crossing experiments and morphological, biological, ecological and molecular analyses. These taxonomic tools showed the closest relationship between T. b. macromelasoma and Triatoma brasiliensis brasiliensis. T. b. macromelasoma is redescribed based on specimens collected in the type locality and specimens from a F1 colony. The complex now comprises T. b. brasiliensis, T. b. macromelasoma, Triatoma melanica, Triatoma juazeirensis and Triatoma sherlocki. An identification key for all members of the complex is presented. This detailed comparative study of the morphological features of T. b. macromelasoma and the remaining members of the complex corroborates results from multidisciplinary analyses, suggesting that the subspecific status is applicable. This subspecies can be distinguished by the following combination of features: a pronotum with 1+1 narrow brownish-yellow stripes on the submedian carinae, not attaining its apex, hemelytra with membrane cells darkened on the central portion and legs with an incomplete brownish-yellow ring on the apical half of the femora. Because the T. brasiliensis complex is of distinct epidemiological importance throughout its geographic distribution, a precise identification of its five members is important for monitoring and controlling actions against Chagas disease transmission.     

First genotyping of Trypanosoma cruzi from naturally infected Triatoma juazeirensis,Triatoma melanica and Triatoma sherlocki from Bahia State,Brazil

Many previous studies have shown a great phylogenetic and biological variability of Trypanosoma cruzi using different molecular and biochemical methods. Populations of T. cruzi were initially clustered into two main lineages called TcI and TcII by the size of the mini‐exon PCR product. In the present study, 33 isolates derived from three triatomine taxa, which belong to the Triatoma brasiliensis species complex (Triatoma juazeirensis, Triatoma melanica and Triatoma sherlocki); collected in three distinct areas of Bahia state were characterized by PCR. The isolates were identified by the size of the mini‐exon gene, 18S rRNA and 24Sα rRNA amplicons. T. cruzi isolates obtained in sylvatic and intradomiciliar ecotopes, derived from T. juazeirensis and T. melanica, were identified as TcI while the parasites originated from T. sherlocki were characterized as TcI and TcII genotypes, respectively. Those species are present in sylvatic ecotopes but are able to infest intradomiciliar areas. Therefore, it would be important to maintain studies in those localities of Bahia and further investigate the possibilities of Chagas disease transmission. Human disease may occur by any T. cruzi genotype and not only by TcII as it is the case in Amazonia.     

Analysis of spermiogenesis like a tool in the study of the triatomines of the Brasiliensis subcomplex

The specific identification and systematic of triatomines have been based fundamentally on morphological observations. These organisms are classified into complexes and specific subcomplexes, principally for morphological parameters and geographical disposition. The use of cytogenetic analyzes has been represented as a tool in systematic and taxonomy of triatomines. Thus, the present work, through the analysis of spermiogenesis, aims to characterize this stage of spermatogenesis in triatomines little studied, and especially to compare it among the species Triatoma lenti and T. sherlocki, to assist in the diagnosis of differentiation of these insects. The presence of the heteropyknotic corpuscle is shown as a diagnostic tool to differentiate T. sherlocki and T. lenti, since it is absent in T. lenti. The analysis of the spermiogenesis in T. sherlocki also allowed us to address morphological differences between elongating cells, which were relatively smaller and more filamentous when compared to T lenti. Furthermore, the flagellum was observed in all stages of cell differentiation and elongation. This structure, which helps in the locomotion of the sperm, is hardly observed in cytogenetic analysis, especially throughout spermiogenesis. Thus, although other comparative approaches should be taken, this paper allowed emphasizing the analysis of spermiogenesis as an important cytotaxonomic tool that assists in the differentiation of morphologically related species, such as T. lenti and T. sherlocki.     

Morphological and morphometric analysis of scutella of six species and two subspecies of Triatoma (Hemiptera: Reduviidae) in Mexico

Morphological characters can be used to distinguish the vast majority of triatomine species, but the existence of high levels of phenotypic plasticity and recently diverged species can lead to erroneous determinations. To approach this problem, we analyzed the male and female morphologies of the scutella of Triatoma barberi, T. dimidiata, T. lecticularia, T. mexicana, T. recurva, T. rubida, and two sub‐species, T. protracta protracta and T. protracta nahuatlae. Scutellum samples were observed by scanning electron microscopy and subjected to morphological analysis and morphometric investigation using a canonical discriminant analysis. The results revealed differences primarily in central depression shape, posterior process, and vestiture. We observed clear dimension‐based differences in scutellum morphometry in all the taxa under study, providing sound evidence for species and subspecies differentiation. On the other hand, there is no difference between sexes in T. lecticularia, T. protracta protracta, and T. protracta nahuatlae. Our methodology can be implemented to differentiate species of the genus Triatoma.     

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.     

Vector capacity of members of Triatoma brasiliensis species complex: The need to extend Chagas disease surveillance to Triatoma melanica

We conducted a lab‐based comparative study on vector capacity features of two species of triatomines: Triatoma brasiliensis and T. melanica. Both are members of the T. brasiliensis species complex. The former is the most important Chagas disease vector in the northeastern region of Brazil. To date, no transmission via T. melanica has been recorded. Immature insects exhibited distinct intermoult periods without a direct relationship to a given species. Females of T. brasiliensis consumed an average of 1.9 times more meals (mean = 12.92 vs 6.63) and survived for a shorter period (mean =330.8 days) than T. melanica (mean = 365.2 days), probably due to the cost of reproduction (all significant at P<0.05). These data support the idea that T. brasiliensis is more adapted to lab conditions and is more able to infest domiciles than T. melanica. We also found significant distinctions in other features between these species, such as the elapsed time without eating before molting, which was higher for the second, third, and fifth nymph stages of T. melanica. Regarding features analyzed related to vector capacity, insects of all life stages of both species were considered competent to transmit Trypanosoma cruzi because they needed many feedings (mean =1.5‐3.82) to moult and because a high proportion (>39%) of insects defecated rapidly (<30 s) after feeding. Overall, results highlight the need to extend vector surveillance to T. melanica.     

Ecotope effect in Triatoma brasiliensis (Hemiptera: Reduviidae) suggests phenotypic plasticity rather than adaptation

Triatoma brasiliensis (Hemiptera: Reduviidae) is an important vector of Chagas' disease in both sylvatic and peridomestic ecotopes. Discriminating between these populations of Triatominae has been proposed as a means of investigating re‐infestation rates of human dwellings. Geometric morphometrics have been widely applied in the study of Triatominae polymorphisms at species and population levels. This study characterizes morphometric differences between sylvatic and peridomestic populations, as well as between sexes in T. brasiliensis specimens from Jaguaruana, Ceará, in northeastern Brazil. No differences in either the shape or size of the cephalic capsule were apparent between sexes or ecotopes. However, the wings showed differentiation in shape and size. Sexual dimorphism was detected, with females presenting significantly higher values and conformations. Size differentiation was also evident, with sylvatic specimens being generally larger than peridomestic examples. These results indicate that differences in the wings of T. brasiliensis may be related to the existence of phenotypic plasticity, and variations in size and shape may be associated with different ecotopes, possibly as a result of conditions in each micro‐habitat, such as temperature, relative humidity, food supply and density.     

Could the bug Triatoma sherlocki be vectoring Chagas disease in small mining communities in Bahia,Brazil?

Searches for Chagas disease vectors were performed at the type locality from which Triatoma sherlocki Papa et al. (Hemiptera: Reduviidae: Triatominae) was described in the municipality of Gentio do Ouro, in the state of Bahia, Brazil, and in a small artisan quarry‐mining community ～13 km distant in a remote area of the same municipality. The latter site represents a new locality record for this species. Adults, nymphs and exuviae of T. sherlocki were found in 21% of human dwellings, indicating that the species is in the process of domiciliation. Prevalence of Trypanosoma cruzi infection in collected bugs was 10.8%. Simple predictive approaches based on environmental similarity were used to identify additional sites likely suitable for this species. The approach successfully predicted an additional five sites for the species in surrounding landscapes. Ecological and entomological indicators were combined to discuss whether this scenario likely represents an isolated case or an emerging public health problem.     

Population structure of a widespread bat (Tadarida brasiliensis) in an island system

Dispersal is a driving factor in the creation and maintenance of biodiversity, yet little is known about the effects of habitat variation and geography on dispersal and population connectivity in most mammalian groups. Bats of the family Molossidae are fast‐flying mammals thought to have potentially high dispersal ability, and recent studies have indicated gene flow across hundreds of kilometers in continental North American populations of the Brazilian free‐tailed bat, Tadarida brasiliensis. We examined the population genetics, phylogeography, and morphology of this species in Florida and across islands of The Bahamas, which are part of an island archipelago in the West Indies. Previous studies indicate that bats in the family Phyllostomidae, which are possibly less mobile than members of the family Molossidae, exhibit population structuring across The Bahamas. We hypothesized that T. brasiliensis would show high population connectivity throughout the islands and that T. brasiliensis would show higher connectivity than two species of phyllostomid bats that have been previously examined in The Bahamas. Contrary to our predictions, T. brasiliensis shows high population structure between two groups of islands in The Bahamas, similar to the structure exhibited by one species of phyllostomid bat. Phylogenetic and morphological analyses suggest that this structure may be the result of ancient divergence between two populations of T. brasiliensis that subsequently came into contact in The Bahamas. Our findings additionally suggest that there may be cryptic species within T. brasiliensis in The Bahamas and the West Indies more broadly.     

Acaulospora brasiliensis comb. nov. and Acaulospora alpina (Glomeromycota) from upland Scotland: morphology, molecular phylogeny and DNA-based detection in roots

Spores of two supposedly arbuscular mycorrhizal fungal species, new to the United Kingdom and recently described as Acaulospora alpina and Ambispora brasiliensis (Glomeromycota), were discovered in soil samples from moorland in upland Scotland. Soil and plant trap pot cultures were established, but attempts to establish these fungi in single-species pot cultures with Plantago lanceolata as host were unsuccessful. Nevertheless, based on a 1.5-kb DNA fragment spanning part of the small subunit rRNA gene, the internal transcribed spacer region and part of the large subunit rRNA gene, both these species could be detected directly in field-sampled roots, together with one uncultured species each of Scutellospora, Rhizophagus (former Glomus group Ab, or ‘Glomus intraradices clade’) and Acaulospora. Whereas A. alpina has characteristic morphological similarities to other species in its genus, A. brasiliensis morphologically has little in common with any other species in Ambispora. The molecular phylogeny, DNA barcoding and morphological evidence clearly place A. brasiliensis in the genus Acaulospora. We therefore rename the species, reported from Brazil and Scotland, as Acaulospora brasiliensis comb. nov., and discuss ecological aspects of the very different environments from which A. brasiliensis and A. alpina have been reported.     

Influence of the intestinal anticoagulant in the feeding performance of triatomine bugs (Hemiptera; Reduviidae)

Triatomines are haematophagous insects in all post-embryonic life stages. They are vectors of Trypanosoma cruzi, the causative agent of Chagas disease. Their vectorial ability is influenced by their feeding performance, which varies greatly amongst species. Recent work showed that inhibition of the coagulation process in the anterior midgut (crop) environment considerably influences the blood meal size. In this work, we performed a comparative study of the level of anticoagulant activity in the saliva and crop contents of three triatomine species - Triatoma infestans, Triatoma brasiliensis and Rhodnius prolixus - and correlated this with their feeding performance on live hosts. Moreover, the feeding parameters on a large diameter vessel influenced by the crop anticoagulants were evaluated in detail. The anticoagulant activity was significantly higher in the crop contents than in salivary glands, varying from 1.6-fold higher for R. prolixus to 70-fold higher for T. brasiliensis. Amongst the species, T. brasiliensis had the lowest crop anticoagulant activity, the lowest concentration of thrombin inhibitor, and took the longest to feed. Triatoma brasiliensis nymphs that had their intestinal anticoagulant (brasiliensin) knocked down by RNA interference had the lowest capacity to maintain cibarial pump frequency at higher levels throughout the feeding process and consequently a lower ingestion rate (mg/min), even when fed under favourable conditions (large diameter vessel). However, the feeding difficulty for brasiliensin knockdown T. brasiliensis nymphs was reversed by treating the host mice with heparin (a potent systemic anticoagulant) before blood feeding. The results indicate that crop anticoagulant activity influences modulation of the blood-pumping frequency to the intestine and significantly affects the feeding efficiency of triatomine spp. on live hosts.     

The cuticular hydrocarbons of the Triatoma sordida species subcomplex (Hemiptera: Reduviidae)

The cuticular hydrocarbons of the Triatoma sordida subcomplex (Hemiptera: Reduviidae: Triatominae) were ana-lysed by gas chromatography and their structures identified by mass spectrometry. They comprised mostly n-alkanes and methyl-branched alkanes with one-four methyl substitutions. n-alkanes consisted of a homologous series from C21-C33 and represented 33-45% of the hydrocarbon fraction; n-C29 was the major component. Methyl-branched alkanes showed alkyl chains from C24-C43. High molecular weight dimethyl and trimethylalkanes (from C35-C39) represented most of the methyl-branched fraction. A few tetramethylalkanes were also detected, comprising mostly even-numbered chains. Several components such as odd-numbered 3-methylalkanes, dimethylalkanes and trimethylalkanes of C37 and C39 showed patterns of variation that allowed the differentiation of the species and populations studied. Triatoma guasayana and Triatoma patagonica showed the most distinct hydrocarbon patterns within the subcomplex. The T. sordida populations from Brazil and Argentina showed significantly different hydrocarbon profiles that posed concerns regarding the homogeneity of the species. Triatoma garciabesi had a more complex hydrocarbon pattern, but it shared some similarity with T. sordida. The quantitative and qualitative variations in the cuticular hydrocarbons may help to elucidate the relationships between species and populations of this insect group.     

Phylogenetic relationships in the Sceloporus variabilis (Squamata: Phrynosomatidae) complex based on three molecular markers,continuous characters and geometric morphometric data

The monophyly of the Sceloporus variabilis group is well established with five species and two species complexes, but phylogenetic relationships within species complexes are still uncertain. We studied 278 specimens in 20 terminals to sample all taxa in the “variabilis group,” including three subspecies in the “variabilis complex,” and two outgroups (Sceloporus grammicus and Sceloporus megalepidurus). We assembled an extensive morphological data set with discrete and continuous characters (distances and scale counts), including geometric morphometric data (landmark coordinates of three shapes), and a three‐marker molecular data set as well (ND4, 12S and RAG1). We conducted parsimony and Bayesian phylogenetic inferences on these data, including several partitioning and weighting schemes. We suggest elevating three subspecies to full species status. Therefore, we recommend recognition of nine species in the “variabilis group.” First, S. variabilis is sister to Sceloporus teapensis. In turn, Sceloporus cozumelae is sister to Sceloporus olloporus. These four species are a monophyletic group, which is sister to Sceloporus smithi. Finally, Sceloporus marmoratus is sister of the clade of five species. The other species in the “variabilis group” (Sceloporus chrysostictus, Sceloporus couchii and Sceloporus parvus) are a paraphyletic grade at the base of the tree. Our analyses reject the existence of the “variabilis complex.” We conducted a parsimony‐based ancestral reconstruction on body size (snout–vent length), femoral pores and dorsal scales and related morphological changes to geographic distribution of the species. Our phylogenetic hypothesis will allow best designs of comparative studies with species in the “variabilis group,” one of the earliest divergent lineages in the genus.     

Genetic and morphological divergence within the Sebastes pachycephalus complex (Scorpaeniformes: Scorpaenidae)

Genetic and morphological divergence among the four subspecies in the Sebastes pachycephalus complex (S. pachycephalus pachycephalus, S. p. nigricans, S. p. nudus and S. p. chalcogrammus) was clarified. Principal coordinate analysis (PCoA) based on AFLP clearly divided 55 specimens of the complex into two groups, the S. p. pachycephalus?CS. p. nigricans group (P-Ni group) and the S. p. nudus?CS. p. chalcogrammus group (Nu-C group), although three specimens occupied intermediate positions. The minimum spanning network (MSN) based on partial sequences of the mitochondrial control region (mtCR) failed to separate either the P-Ni and Nu-C groups or the four subspecies into distinct clades, although restricted gene flow and genetic differentiation between the former were indicated by the F _ST estimation. Differences in morphological characters, including counts of pectoral fin rays and counts of dorsal fin spines lacking basal scales, were also evident between the two groups. However, little or no genetic or morphological difference was found between the two subspecies within each group. It was concluded that the P-Ni and Nu-C groups of the S. pachycephalus complex actually represent two different species, which is further supported by their sympatric distribution. Differences in dorsal body coloration and the presence or absence of brown spots on the ventral surface, which were formerly used to discriminate between four ??subspecies,?? may simply represent intraspecific variation. The three specimens occupying intermediate positions in the AFLP PCoA also occupied equivocal positions between the two species in the principal component analysis (PCA) based on morphometric characters, suggesting that they were hybrids between the two species. The star-shaped MSN of mtCR, which lacks distinct clades representing the two species, may be due to not only interspecific hybridization but also the sharing of ancestral haplotypes.     

Cylicocyclus asini n. sp. (Nematoda: Cyathostominae) from donkeys Equus asinus in South Africa

Cylicocyclus asini n. sp. is described from the ventral colons of seven domesticated donkeys Equus asinus in South Africa. The specimens are smaller than 11 members of the genus Cylicocyclus, which currently comprises 10 recognised species, one recognised subspecies and two species inquirendae, but is similar in size to the two smaller species, C. ashworthi and C. leptostomum. Additionally, the small size of its buccal capsule places it within the so-called small buccal capsule group, namely C. ashworthi, C. leptostomum, C. nassatus, C. radiatus and C. triramosus. The shape of the oesophagus at the oesophago-intestinal junction is elongate, similar to that of C. leptostomum. The female specimens have a tail length shorter than the vulva to anus distance, similar to three of the smaller Cylicocyclus species, namely C. leptostomum, C. radiatus and C. triramosus. The new species is distinguished from the smaller members of the genus by the presence of a `club-foot' posterior in the female specimens, 40–46 elements in the external leaf-crown and a deep division of the dorsal ray which extends beyond the origin of the externodorsal ray. Similarly, the new species can be separated from the two other members in the genus with a `club-foot' posterior in the females (C. auriculatus and C. gyalocephaloides) by its body length (6.6–8.47 mm) and the presence of a nipple-like dorsal gutter.     

External female genitalia of six species of the genus Meccus (Hemiptera: Reduviidae: Triatominae)

Triatomine classification is based on morphological characteristics. Studies have been conducted to improve their identification by observing many characteristics. However, there are problems of differentiating among highly interrelated species and new criteria are required. The purpose of this study was to determine the morphological differences in the external female genitalia of M. pallidipennis, M. longipennis, M. picturatus, M. bassolsae, M. mazzottii, and M. phyllosomus in order to distinguish among species using scanning electron microscopy. Observations were made of the dorsal, posterior, lateral, and ventral views of the female external genitalia for each species. In the six species we studied, relevant differences were observed in the dorsal view of the X segment, as well as the IX, VIII, and VII tergites. In the posterior and lateral view, the most visible differences were registered in the gonocoxite size of the segments VIII, IX, and X. Finally, in the ventral view of the VII sternite, differences among species were observed in the size of the inflection in the top and upper corner. Our results show that it was possible to differentiate among the triatominae species for each of the four views by using a scanning electron microscope to analyze morphological characteristics of the VII, VIII, IX, and X abdomen segments.     

On the variability of fire-bellied toads in the Far East

We studied external morphology and molecular genetics of two forms of the oriental fire-bellied toad, described as separate subspecies Bombina orientalis silvatica Korotkov, 1972, and Bombina orientalis practicola Korotkov, 1972. We have found significant differences between them in body size, coloration and some morphometric parameters but not in genetics. Taking into account their geographical separation, one can consider them as forest and meadow ecological morphs but not different taxa. Therefore, the names Bombina orientalis practicola Korotkov, 1972, and B. orientalis silvatica Korotkov, 1972, should be considered as junior synonyms of the name Bombina orientalis (Boulenger, 1980). We suppose that these morphs have been isolated from each other recently, and their morphological differences at genetic similarity are related to high plasticity of the species, when ecologically determined morphological differentiation precedes genetic differentiation.     

The biology of three Mexican-American species of Triatominae (Hemiptera: Reduviidae): Triatoma recurva, Triatoma protracta and Triatoma rubida

The values of biological parameters related to hatching, lifespan, the number of blood meals between moults, mortality, time lapse before the beginning of feeding, feeding time and defecation delay for each instar of three Mexican-American species of Triatominae, Triatoma recurva, Triatoma protracta (former subspecies protracta) and Triatoma rubida (former subspecies uhleri), were evaluated and compared. No significant (p > 0.05) differences were recorded among the three species with respect to the average time required to hatch. This time was approximately 19 days. The average egg-to-adult development time was significantly (p < 0.05) shorter for T. rubida. The number of blood meals at each nymphal instar varied from one-five for each species. The mortality rates were higher for the first-instar nymphs of the three species studied. The mean time lapse before the beginning of feeding was between 0.3-3 min for most nymphs of all instars of each species studied. The mean feeding time was the longest for T. recurva, followed by T. protracta. The defecation delay was less than 10 min for T. recurva and T. rubida. Given these results, only T. rubida should be considered an important potential vector of Trypanosoma cruzi transmission to humans in areas of Mexico where these species exist, whereas T. recurva and T. protracta would be of secondary importance.