Haplodontium zangii X.R.Wang & J.C.Zhao,sp. nov. (Bryaceae,Bryopsida) from Xizang,China

Haplodontium zangii X.R.Wang &; J.C.Zhao, a new moss species from Xizang, China, is described and illustrated. Previously, specimens of H. zangii have been identified as Mielichhoferia himalayana Mitt. However, H. zangii is distinctly different from M. himalayana in having excurrent costae with short awns (vs long denticulate awns), short-pyriform capsules, 0.8–1?mm (vs long-pyriform capsules, 2.5–3?mm), and densely papillose exostome teeth (vs smooth or vertically striped exostome teeth). Haplodontium zangii is similar to H. macrocarpum (Hook.) J.R.Spence, which was traditionally placed in Mielichhoferia Nees &; Hornsch. as M. macrocarpa (Hook.) Bruch &; Schimp. The main differences between H. zangii and H. macrocarpum are in the morphology of the leaves, capsules, guide cells, and stomata. Mielichhoferia himalayana and another Chinese species of Mielichhoferia, M. sinensis Dix., are also transferred to Haplodontium Hampe, a new genus in the bryoflora of China, as H. himalayanum (Mitt.) X.R.Wang &; J.C.Zhao and H. sinensis (Dix.) X.R.Wang &; J.C.Zhao. A morphological comparison and a key to the three species of Haplodontium in China as well as to H. macrocarpum, a species that is likely to be found in China, are provided.     

Isozyme variability and differentiation between Rhodnius prolixus, R.robustus and R.pictipes, vectors of Chagas disease in Venezuela

Enzyme polymorphism in triatomine bugs of the genus Rhodnius (Hemiptera: Reduviidae), vectors of Chagas disease, is analysed using both starch and polyacrylamide gel electrophoresis. Out of forty-five enzymes assayed, the electromorphs of seventeen of them: AO, CA, DIA, ES, ES-A, FH, GPD, G6PD, GPI, MDH, ME, 6PGD, PGM, ACON, ACPH, LAP and SOD, involving twenty-two putative structural loci, were scorable. These gene-enzyme systems were therefore selected for routine characterization of R.prolixus Stal adults from different strains. The first thirteen enzymes, involving sixteen structural loci, were also analysed in first instar nymphs of the three species, R.prolixus, R.robustus Larrousse and R.pictipes Stal. Allelic frequencies were calculated for three R.prolixus strains: three to five loci appeared to be polymorphic. The proportion of polymorphic loci (22%) and the average heterozygosity (0.06) indicated low genetic variability, with significant differences between the strains at individual loci. Rhodnius prolixus and R.robustus were found to have identical isozymic patterns. R.pictipes was genetically well differentiated, with twelve diagnostic loci.     

Genome constitutions of Roegneriaalashanica,R. elytrigioides,R. magnicaespes and R. grandis (Poaceae: Triticeae) via genomic in‐situ hybridization

The genomic constitutions of Roegneria alashanica, R. elytrigioides, R. magnicaespes and R. grandis were studied using GISH. DNA of Pseudoroegneria spicata (St), P. libanotica (St), P. stipifolia (St), R. ciliaris (StY), Lopophyllum elongatum (E^e), Agropyron cristatum (P) and Hordeum bogdanii (H) were used for probing, respectively. The results indicated that: 1) R. alashanica and R. magnicaespes contained one St genome, the other genome was unidentified, however, it was not an E, P, H or Y genome; 2) R. elytrigioides contained two St genomes and should on this basis be included in Pseudoroegneria; 3) R. grandis contained an St and an St^g genome. The St^g genome is suggested to be a modified form of the St genome of Pseudoroegneria and to be homoeologous with the Y genome. It might be an intermediate type between the St and Y genomes. Therefore, R. elytrigioides should be treated as Pseudoroegneriaelytrigioides.Roegneria alashanica, R. magnicaespes and R. grandis does apparently not belong in the genus Roegneria but further studies are needed to establish their correct taxonomic position.     

Analyzing the Impact of Ramalina digitellata,R. fastigiata,R. fraxinea,and R. polymorpha's Usnic Acid Concentration on Antioxidant,DNA-Protective,Antimicrobial, and Cytotoxic Properties

The present study is focused on the antimicrobial, antioxidant, cytotoxic, and DNA protective effects of methanol extract obtained from R. digitellata, R. fastigiata, R. fraxinea, and R. polymorpha species that are distributed in Turkey. The highest total phenol content was determined in R. digitellata (144.6 mgGAE/g_extract), and the highest total amount of flavonoids was found in R. fastigiata (20.40 mgGAE/g_extract). The content of usnic acid was determined by High-Performance Liquid Chromatography (HPLC) and the highest amount was found in R. digitellata. DPPH (1,1-diphenyl-2-picrylhydrazyl) and ABTS [2,2’-azinobis(3-ethylbenzathiazoline-6-sulfonic acid)] radical scavenging methods were used for antioxidant activity. R. fraxinea showed the highest DPPH⋅ and ABTS⁺⋅ scavenging activity. In addition, the DNA protective effect was investigated using pBR322 plasmid DNA, and; all studied species were found to have DNA protective effects. The antibacterial activity was investigated using the disc diffusion method, and the R. digitellata methanol extract showed the best results with a 12.35 mm zone on Proteus mirabilis. On the human lung cancer (A549) and breast cancer (MDA-MB-231) cell lines, cytotoxic activity was assessed using an MTT assay. All lichen extracts were found to have a significant cytotoxic effect on both cancer cell lines at 1000 μg/mL concentration. These results suggest that Ramalina species may be potential candidates for developing new phytopharmaceuticals and functional components.     

Meiotic differences between three triatomine species (Hemiptera,Reduviidae)

We have found the following differences in the male meiosis among three triatomine species: (1) The three largest autosomal bivalents ofTriatoma infestans are heterochromatic.Rhodnius prolixus has two autosomal bivalents with heterochromatic blocks.Triatoma rubrovaria does not show any heteropycnotic autosomes. (2) Sex chromosomes inT. infestans form a chromocenter. At early prophase terminal associations are seen between sex chromosomes inT. rubrovaria, and they maintain a close association until diakinesis. An intimate association between the X and Y chromosomes is observed during early prophase inR. prolixus, but a distant association is maintained by the sex chromosomes at diffuse and diplotene stages in this species. (3) Polyploid nuclei of the nutritive cells are quite distinct. Numerous chromocenters of different shapes and sized are seen in those ofT. infestans. InT. rubrovaria one chromocenter having two positively heteropycnotic elements is observed surrounded by homogeneous chromatin. Only one compact chromocenter is found amongst unevenly distributed chromatin, inR. prolixus.     

Transepithelial transport of salicylate by the Malpighian tubules of insects from different orders

The organic anion salicylate is a plant secondary metabolite that protects plants against phytophagous insects. In this study, a combination of salicylate-selective microelectrodes and a radioisotope tracer technique was used to study the transepithelial transport of salicylate by the Malpighian tubules of 10 species of insects from five orders. Our results show that salicylate is transported into the lumen of the Malpighian tubules in all the species evaluated, except Rhodnius prolixus. The transepithelial transport of salicylate by the Malpighian tubules of Drosophila simulans, Drosophila erecta, Drosophila sechellia, and Acheta domesticus was saturable, Na⁺-dependent and inhibited by α-cyano-4-hydroxycinnamic acid. This transport system resembles that previously found in tubules of Drosophila melanogaster. In contrast, transepithelial transport of salicylate by Malpighian tubules of Tenebrio molitor, Plagiodera versicolora, Aedes aegypti, and Trichoplusia ni was unaffected by Na⁺-free bathing saline. The presence of both salicylate and salicylate metabolites in the secreted fluid samples from the Malpighian tubules of A. domesticus, R. prolixus, T. molitor, and T. ni indicates that insect Malpighian tubules may both transport and metabolize salicylate. The highest capacities to rid the hemolymph of salicylate were found in T. molitor, P. versicolora and Drosphila spp. Our results suggest that transport of salicylate by the Malpighian tubules might contribute to elimination of this organic anion from the hemolymph, particularly in some species that encounter high levels of organic anion in the diet.     

The formation of the antenna sensory apparatus in some bug (Heteroptera) species in the course of their postembryonic development

In the antenna sensory apparatus of bugs Coreus marginatus, Cimex lectularius, and Rhodnius prolixus sensilla of the four main types are identified: chaetica, trichodea, basiconica, and coeloconica. Chaetoid sensilla are differentiated into two subtypes: sensilla with cogged cuticles and those with smooth ones; trichoid sensilla were divided into long pointed and short ones with blunt tips. In larvae and adults of R. prolixus trichobothria (long filiform hairs) were found on the medial side of pedicellum. The postembryonic changes in the quantitative and qualitative composition of the antenna sensory apparatus were assessed using biometric analysis. The greatest increase of sensory organs was observed upon the nymphal ecdysis from the 5th instar to adult.     

The neuropeptidome of Rhodnius prolixus brain

We show a sensitive and straightforward off‐line nano‐LC‐MALDI‐MS/MS workflow that allowed the first comprehensive neuropeptidomic analysis of an insect disease vector. This approach was applied to identify neuropeptides in the brain of Rhodnius prolixus, a vector of Chagas disease. This work will contribute to the annotation of genes in the ongoing R. prolixus genome sequence project. Peptides were identified by de novo sequencing and comparisons to known neuropeptides from different organisms by database search. By these means, we were able to identify 42 novel neuropeptides from R. prolixus. The peptides were classified as extended FMRF‐amide‐related peptides, sulfakinins, myosuppressins, short neuropeptide F, long neuropeptide F, SIF‐amide‐related peptides, tachykinins, orcokinins, allatostatins, allatotropins, calcitonin‐like diuretic hormones, corazonin, and pyrokinin. Some of them were detected in multiple isoforms and/or truncated fragments. Interestingly, some of the R. prolixus peptides, as myosuppressin and sulfakinins, are unique in their characteristic C‐terminal domain among insect neuropeptides identified so far.     

Historical and recent evidence for close relationships among Rickettsia parkeri,R. conorii,R. africae,and R. sibirica: Implications for rickettsial taxonomy

Rickettsia parkeri, a member of the spotted fever group rickettsias, was first described in 1939 and was thought to be non‐pathogenic until recently, when it was found to cause a spotted fever‐like illness in humans and areas of necrosis (eschars) at the sites of tick bites. Accordingly, there is currently much interest in this emerging pathogen. In this study, all published articles concerning R. parkeri were reviewed and analyzed for evidence of relatedness among this agent and other spotted fever group (SFG) rickettsiae which also produce similar clinical syndromes and/or eschars, including R. conorii, R. africae, and R. sibirica. A synthesis of the historical (antigenic) and recent (molecular) data supporting a phylogenetic sub‐grouping of these SFG organisms is presented and comments are offered about the taxonomy of rickettsial organisms in general, and R. parkeri in particular.     

Rhaponticoidesaytachii sp. nov. (Asteraceae) from south Anatolia,Turkey

A new species of Rhaponticoides Vaill., Rhaponticoidesaytachii Y. Ba?c?, Do?u & Dinç sp. nov. collected by the authors from south Anatolia, is described and illustrated. The new species is restricted to deprived Quercus forest openness around Dumlugöze‐Sar?veliler (C4 Karaman). Diagnostic morphological characters from R. amplifolia (Boiss. & Heldr.) M.V. Agab. & Greuter are discussed. The ecology, biogeography and conservation status of the new species is also presented.     

Allozyme variation among Rana balcanica,R. levantina,and R. ridibunda (Amphibia: Anura)1

The allozyme variation among water frogs of the species R. balcanica, R. levantina and R. ridibunda, all formerly considered as one species (R. ridibunda Pallas, 1771), was studied using horizontal starch gel electrophoresis. Blood samples (N # 63) of frogs were collected from five populations in Greece and Israel. Samples (N = 9) of the hybrid frog R. esculenta collected from a locality in Germany were used as an outgroup for phylogenetic analyses. Fifteen enzymes controlled by twenty presumptive loci were identified. Thirteen loci were polymorphic within or among the studied populations. Genetic differentiation among the species was considerably greater than among populations of the same species. Even at Nestos River where R. ridibunda and R. balcanica occur in the same habitats, individuals could be assigned to either species due to characteristic differences of the genotypes (GPI1). This indicates reproductive isolation among these species. The reconstruction of phylogenetic relationships among the three species based on the allozyme data corroborated the model presented on the basis of bioacoustic data: R. ridibunda and R. balcanica (Nei's genetic distance D = 0.0820) are sibling species pertaining to an Eurasian lineage, whereas R. levantina (distance to the European species D = 0.1780 - 0.1955) together with R. perezi represent an independent afroasian lineage.     

Infestation of arboreal nests of coatis by triatomine species,vectors of Trypanosoma cruzi,in a large Neotropical wetland

The coati (Nasua nasua, Carnivora) is a medium‐sized mammal common in the Pantanal of Brazil. Unlike most mammals, coatis construct arboreal nests used for resting and reproduction. In this region, the coati is an important host of Trypanosoma cruzi, the causative agent of Chagas disease. There are two possible routes through coatis can be infected by T. cruzi: the oral route or the vectorial route. However, the relative importance of each of these routes in the infection of coatis and its role in the sylvatic cycle of the parasite are unknown. Our objectives were to investigate: (i) whether coati nests were infested by triatomine bugs, (ii) what species were frequent in the nests, (iii) whether the triatomines in nests were infected by T. cruzi, and (iv) what were the food resources of these triatomines. Eight of the 24 nests sampled were infested with triatomines, a total of 37 specimens of at least two species (Rhodnius stali and Triatoma sordida). In one nest, R. stali and T. sordida co‐occurred and both fed on multiple resources, including coatis. This is the first report of triatomines occurring in arboreal nests of coatis. The co‐occurrence of two different genera of triatomine vectors and coatis within the limited space of the coati nests provide multiple opportunities for the exchange of the protozoan parasite through both the vectorial and oral transmission routes.     

Sulfated glycosaminoglycans in two hematophagous arthropod vectors of Chagas disease, Triatoma brasiliensis and Rhodnius prolixus (Hemiptera: Reduviidae)

The characterization of sulfated glycosaminoglycans (GAGs) in hematophagous arthropod vectors in general has been limited, with the exception of the studies in the triatomine Rhodnius prolixus. Heparan sulfate (HS) and chondroitin sulfate (CS) were previously identified and structurally characterized in extracts of whole bodies of fourth instar larvae of R. prolixus. Recently, we showed the expression of these two sulfated GAGs in specific body tissues of adult males and females and in embryos of R. prolixus. In the present work, we identified and compared the sulfated GAG composition in specific tissues of adult insects and in embryos of another triatomine species, Triatoma brasiliensis. Sulfated GAGs were isolated from the fat body, intestinal tract, and the reproductive tracts of adult insects and from embryos. Only HS and CS were found in the tissues analyzed. The present results extend the initial observations on the sulfated GAG composition in R. prolixus by showing that these molecules are widely distributed among internal organs of triatomines. These observations may be useful for future investigations aiming to evaluate the possible implication of these compounds in physiological events that take place in a specific organ(s) in these insects.     

Comparative proteomic analysis of the saliva of the Rhodnius prolixus,Triatoma lecticularia and Panstrongylus herreri triatomines reveals a high interespecific functional biodiversity

Triatomines are hematophagous arthropods that transmit Trypanosoma cruzi and Trypanosoma rangeli. Feeding behavior and pathogen transmission is known to vary between the different species, and this characteristic is directly or indirectly dependent on the bioactive molecules of the saliva that facilitate the vector–host–parasite interaction. Here, we identify, characterize and compare the sialoproteomic (from the Greek sialo: saliva) repertoire of important species of the main triatomine genera in the Americas (Rhodnius prolixus, Triatoma lecticularia and Panstrongylus herreri) to better explain this interaction through two-dimensional electrophoresis and mass spectrometry. We identified 221 proteins, 69 from R. prolixus, 100 from T. lecticularia and 52 from P. herreri. We identified high abundance molecules with a great potential to modulate host defenses and homeostasis, highlighting Nitrophorin-4 (28.7%), Salivary lipocalin-5 (65.2%) and Putative triabin (20.5%) in R. prolixus, T. lecticularia and P. herreri, respectively. We also observed that only a single hypothetical protein is shared among three species, which was not functionally categorized. This study corroborates previous findings with R. prolixus, increasing the knowledge about this species with relevant proteomic information and comparisons with the other two targets of the study, T. lecticularia and P. herreri, for which no studies are available from a proteomics perspective.     

Dislodgement effect of natural semiochemicals released by disturbed triatomines: a possible alternative monitoring tool

The quick detection of domestic and peridomestic triatomines in their environments becomes difficult without the use of dislodgement substances that flush them out from their shelters. At present, tetramethrin 0.2% is being widely used in control programs. Although it is an efficient dislodging agent, its toxicity might affect the health of captured triatomines, of other insects and, to a lesser extent, of other animals, including humans. Here, we tested if semiochemicals released by disturbed adults of Triatoma infestans and/or Rhodnius prolixus can make larvae of the same species exit from their refuges. In a walking olfactometer we found that: 1) larvae of T. infestans were repelled by the odors released by disturbed adults of their own species and of R. prolixus, 2) larvae of R. prolixus did not change their behavior in the presence of odors released by adults of both species, and 3) activity levels were not modulated by these odors in any of both species. Besides, in pseudo‐natural conditions we found an increased flushing‐out activity of larvae of T. infestans when their shelters were sprayed with isobutyric acid or 3‐pentanol, and of larvae of R. prolixus when sprayed with 3‐methyl1butanol. We succeeded in this work to dislodge larvae of triatomines from artificial shelters using natural volatile compounds, allowing the capture of live bugs for further investigations (e.g., xenodiagnosis or genetic studies) and favoring ecological aspects (e.g., minimizing environmental insecticide‐contamination and non‐targeted mortality).     

Female metasternal gland odours mediate male aggregation in Rhodnius prolixus, a triatomid bug

We investigated whether metasternal gland (MG) products mediate male aggregation during mating in Rhodnius prolixus (Heteroptera: Reduviidae) Stål, 1859, through selective occlusion of female MGs. Pairs of insects presenting intact MGs mated and other males aggregated around them confirming conclusions from previous studies. However, when female MGs were occluded, males did not aggregate. Conversely, the occlusion of male MGs did not affect aggregation of males. The present experiments demonstrated that the products emitted by the MGs of a mating R. prolixus female induce other males to aggregate around the mating pair. Aggregation behaviour of males probably results from the eventual release of volatiles from the female's MGs. We reinforce the hypothesis suggesting that the compounds produced by the MGs of R. prolixus act as sexual communication signals for this species.     

Molecular genetics reveal that silvatic Rhodnius prolixus do colonise rural houses

Background

Rhodnius prolixus is the main vector of Chagas disease in Venezuela. Here, domestic infestations of poor quality rural housing have persisted despite four decades of vector control. This is in contrast to the Southern Cone region of South America, where the main vector, Triatoma infestans, has been eliminated over large areas. The repeated colonisation of houses by silvatic populations of R. prolixus potentially explains the control difficulties. However, controversy surrounds the existence of silvatic R. prolixus: it has been suggested that all silvatic populations are in fact Rhodnius robustus, a related species of minor epidemiological importance. Here we investigate, by direct sequencing (mtcytb, D2) and by microsatellite analysis, 1) the identity of silvatic Rhodnius and 2) whether silvatic populations of Rhodnius are isolated from domestic populations.

Methods and Findings

Direct sequencing confirmed the presence of R. prolixus in palms and that silvatic bugs can colonise houses, with house and palm specimens sharing seven cytb haplotypes. Additionally, mitochondrial introgression was detected between R. robustus and R. prolixus, indicating a previous hybridisation event. The use of ten polymorphic microsatellite loci revealed a lack of genetic structure between silvatic and domestic ecotopes (non-significant F_ST values), which is indicative of unrestricted gene flow.

Conclusions

Our analyses demonstrate that silvatic R. prolixus presents an unquestionable threat to the control of Chagas disease in Venezuela. The design of improved control strategies is essential for successful long term control and could include modified spraying and surveillance practices, together with housing improvements.     

Gymnosoma rotundatum (Diptera: Tachinidae) attracted to the aggregation pheromone of Plautia stali (Hemiptera: Pentatomidae)

The brown-winged green stink bug, Plautia stali Scott (Hemiptera: Pentatomidae), is an important fruit-piercing bug. A monitoring study on the seasonal density change of P. stali using traps baited with methyl (E,E,Z)-2,4,6-decatrienoate, the aggregation pheromone of the P. stali, resulted in the first capture of adult tachinid flies, Gymnosoma rotundatum (Diptera: Tachinidae), in Korea. The parasitoid fly may worth studying as a biological control agent in the management of P. stali.     

Neutral sterols of representatives of two groups of hemiptera and their correlation to ecdysteroid content

The sterols of four species of Pentatomomorpha—Oncopeltus fasciatus (Dallas), Nezara viridula (L.), Dysdercus cingulatus (F.), and Podisus maculiventris (Say)—and threé species of Cimicomorpha—Rhodnius prolixus Stal, Arilus cristatus (L), and Cimex lectularius (L.)—were isolated and examined in order to compare neutral sterol utilization and content with the known ecdysteroids of these species. In the phytophagous Pentatomomorpha, O. fasciatus, N. viridula, and D. cingulatus, the low content of cholesterol and the high content of C₂₈ and C₂₉ phytosterols (< 1% and > 99% of the tissue sterols, respectively) indicated that these species are unable to dealkylate the C-24 position of sterols. These insects appear to have adapted to the challenge of both insufficient dietary cholesterol and inability to dealkylate phytosterols by evolving the ability to produce a C₂₈ ecdysteroid (makisterone A). The secondarily predacious P. maculiventris has adequate cholesterol available for C₂₇ ecdysteroid production, but appears to have retained the ecdysteroid biosynthetic pathways of its phytophagous ancestors because it produces a C₂₈ ecdysteroid. Cholesterol was the major sterol of each of the three species of Cimicomorpha, and these insects are only able to produce C₂₇ ecdysteroids.