Spatio-Temporal Tracking and Phylodynamics of an Urban Dengue 3 Outbreak in Sao Paulo, Brazil

The dengue virus has a single-stranded positive-sense RNA genome of similar to 10.700 nucleotides with a single open reading frame that encodes three structural (C, prM, and E) and seven nonstructural (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5) proteins. It possesses four antigenically distinct serotypes (DENV 1-4). Many phylogenetic studies address particularities of the different serotypes using convenience samples that are not conducive to a spatio-temporal analysis in a single urban setting. We describe the pattern of spread of distinct lineages of DENV-3 circulating in Sao Jose do Rio Preto, Brazil, during 2006. Blood samples from patients presenting dengue-like symptoms were collected for DENV testing. We performed M-N-PCR using primers based on NS5 for virus detection and identification. The fragments were purified from PCR mixtures and sequenced. The positive dengue cases were geo-coded. To type the sequenced samples, 52 reference sequences were aligned. The dataset generated was used for iterative phylogenetic reconstruction with the maximum likelihood criterion. The best demographic model, the rate of growth, rate of evolutionary change, and Time to Most Recent Common Ancestor (TMRCA) were estimated. The basic reproductive rate during the epidemics was estimated. We obtained sequences from 82 patients among 174 blood samples. We were able to geo-code 46 sequences. The alignment generated a 399-nucleotide-long dataset with 134 taxa. The phylogenetic analysis indicated that all samples were of DENV-3 and related to strains circulating on the isle of Martinique in 2000-2001. Sixty DENV-3 from Sao Jose do Rio Preto formed a monophyletic group (lineage 1), closely related to the remaining 22 isolates (lineage 2). We assumed that these lineages appeared before 2006 in different occasions. By transforming the inferred exponential growth rates into the basic reproductive rate, we obtained values for lineage 1 of R(0) = 1.53 and values for lineage 2 of R(0) = 1.13. Under the exponential model, TMRCA of lineage 1 dated 1 year and lineage 2 dated 3.4 years before the last sampling. The possibility of inferring the spatio-temporal dynamics from genetic data has been generally little explored, and it may shed light on DENV circulation. The use of both geographic and temporally structured phylogenetic data provided a detailed view on the spread of at least two dengue viral strains in a populated urban area.

An assessment of population structure in eight breeds of cattle using a whole genome SNP panel

Background: Analyses of population structure and breed diversity have provided insight into the origin and evolution of cattle. Previously, these studies have used a low density of microsatellite markers, however, with the large number of single nucleotide polymorphism markers that are now available, it is possible to perform genome wide population genetic analyses in cattle. In this study, we used a high-density panel of SNP markers to examine population structure and diversity among eight cattle breeds sampled from Bos indicus and Bos taurus. Results: Two thousand six hundred and forty one single nucleotide polymorphisms ( SNPs) spanning all of the bovine autosomal genome were genotyped in Angus, Brahman, Charolais, Dutch Black and White Dairy, Holstein, Japanese Black, Limousin and Nelore cattle. Population structure was examined using the linkage model in the program STRUCTURE and Fst estimates were used to construct a neighbor-joining tree to represent the phylogenetic relationship among these breeds. Conclusion: The whole-genome SNP panel identified several levels of population substructure in the set of examined cattle breeds. The greatest level of genetic differentiation was detected between the Bos taurus and Bos indicus breeds. When the Bos indicus breeds were excluded from the analysis, genetic differences among beef versus dairy and European versus Asian breeds were detected among the Bos taurus breeds. Exploration of the number of SNP loci required to differentiate between breeds showed that for 100 SNP loci, individuals could only be correctly clustered into breeds 50% of the time, thus a large number of SNP markers are required to replace the 30 microsatellite markers that are currently commonly used in genetic diversity studies.

Ovariole structure and oogenesis in queens and workers of the stingless bee Melipona quadrifasciata (Hymenoptera: Apidae, Meliponini) kept under different social conditions

The high variability in the reproductive biology of stingless bees makes them very amenable for comparative studies with other eusocial bee taxa. We investigated the structural organization of the ovaries of Melipona quadrifasciata queens and workers kept under different social conditions by analyzing their general histology, mitotic activity, and microfilament organization. The overall dynamics of ovarian activity were similar in the two castes, and at emergence their ovarioles contained a previtellogenic follicle. Stingless bees and honey bees differ in the structural organization in the lower germarium, but they have in common synchronized mitotic activity and putative germ line stem cells in the terminal filament. Unlike honey bees, stingless bee workers lay trophic eggs in addition to reproductive eggs. The overall similarities in oogenesis between the two taxa suggest that the decision to form trophic eggs should only occur in the late stages of oogenesis.

FBXO25-associated nuclear domains: A novel subnuclear structure

Skp1, Cul1, Rbx1, and the FBXO25 protein form a functional ubiquitin ligase complex. Here, we investigate the cellular distribution of FBXO25 and its colocalization with some nuclear proteins by using immunochemical and biochemical approaches. FBXO25 was monitored with affinity-purified antibodies raised against the recombinant fragment spanning residues 2-62 of the FBXO25 sequence. FBXO25 protein was expressed in all mouse tissues tested except striated muscle, as indicated by immunoblot analysis. Confocal analysis revealed that the endogenous FBXO25 was partially concentrated in a novel dot-like nuclear domain that is distinct from clastosomes and other well-characterized structures. These nuclear compartments contain a high concentration of ubiquitin conjugates and at least two other components of the ubiquitin-proteasome system: 20S proteasome and Skp1. We propose to name these compartments FBXO25-associated nuclear domains. Interestingly, inhibition of transcription by actinomycin D or heat-shock treatment drastically affected the nuclear organization of FBXO25-containing structures, indicating that they are dynamic compartments influenced by the transcriptional activity of the cell. Also, we present evidences that an FBXO25-dependent ubiquitin ligase activity prevents aggregation of recombinant polyglutamine-containing huntingtin protein in the nucleus of human embryonic kidney 293 cells, suggesting that this protein can be a target for the nuclear FBXO25 mediated ubiquitination.

Observations on Antricola ticks: Small nymphs feed on mammalian hosts and have a salivary gland structure similar to ixodid ticks

Ticks use bloodmeals as a Source of nutrients and energy to molt and survive until the next meal and to oviposit, in the case of females. However, only the larvae of some tick species are known to feed upon bats females are obligatorily autogenous, and nymphal stages are believed to not feed. We investigated the presence of blood ill a natural population of nymphal Antricola delacruzi ticks collected from bat guano; their ability to feed upon laboratory hosts: and the microscopic structure of both salivary glands and gut. DNA amplification of gut contents of freshly collected material was positive for a mammal in 4 of 11 first instar nymphs, but we were unsuccesful in the amplification of host bloodmeal DNA from late instar nymphs. All early nymphal stages (n = 10) fed oil rabbits. and host DNA was detected and sequenced from gut contents. However, all the large nymphs (n = 10) rejected feeding, and host DNA remained undetected in these ticks. All stages of A. delacruzi have salivary glands similar in morphology to the ixodid agranular Type I salivary gland acini and to granular Type II or Type B acini. All stages of A. delacruzi had a similar gut structure. consisting of digestive cells in the basal portion that contained hematin granules. Neither regenerative nor secretory cell traces were observed in the sections Of gut.

Cost effectiveness of OptiMal (R) rapid diagnostic test for malaria in remote areas of the Amazon Region, Brazil

Background: In areas with limited structure in place for microscopy diagnosis, rapid diagnostic tests (RDT) have been demonstrated to be effective. Method: The cost-effectiveness of the Optimal (R) and thick smear microscopy was estimated and compared. Data were collected on remote areas of 12 municipalities in the Brazilian Amazon. Data sources included the National Malaria Control Programme of the Ministry of Health, the National Healthcare System reimbursement table, hospitalization records, primary data collected from the municipalities, and scientific literature. The perspective was that of the Brazilian public health system, the analytical horizon was from the start of fever until the diagnostic results provided to patient and the temporal reference was that of year 2006. The results were expressed in costs per adequately diagnosed cases in 2006 U. S. dollars. Sensitivity analysis was performed considering key model parameters. Results: In the case base scenario, considering 92% and 95% sensitivity for thick smear microscopy to Plasmodium falciparum and Plasmodium vivax, respectively, and 100% specificity for both species, thick smear microscopy is more costly and more effective, with an incremental cost estimated at US$ 549.9 per adequately diagnosed case. In sensitivity analysis, when sensitivity and specificity of microscopy for P. vivax were 0.90 and 0.98, respectively, and when its sensitivity for P. falciparum was 0.83, the RDT was more cost-effective than microscopy. Conclusion: Microscopy is more cost-effective than OptiMal (R) in these remote areas if high accuracy of microscopy is maintained in the field. Decision regarding use of rapid tests for diagnosis of malaria in these areas depends on current microscopy accuracy in the field.

The structure and dynamics of Abell 1942

Aims. We present a dynamical analysis of the galaxy cluster Abell 1942 based on a set of 128 velocities obtained at the European Southern Observatory. Methods. Data on individual galaxies are presented and the accuracy of the determined velocities as some properties of the cluster are discussed. We have also made use of publicly available Chandra X-ray data. Results. We obtained an improved mean redshift value z = 0.22513 +/- 0.0008 and velocity dispersion sigma = 908(139)(+147) km s(-1). Our analysis indicates that inside a radius of similar to 1.5 h(70)(-1) Mpc (similar to 7 arcmin) the cluster is well relaxed, without any remarkable features and the X-ray emission traces the galaxy distribution fairly well. Two possible optical substructures are seen at similar to 5 arcmin from the centre in the northwest and the southwest directions, but are not confirmed by the velocity field. These clumps are, however, kinematically bound to the main structure of Abell 1942. X-ray spectroscopic analysis of Chandra data resulted in a temperature kT = 5.5+/-0.5 keV and metal abundance Z = 0.33 +/- 0.15 Z(circle dot). The velocity dispersion corresponding to this temperature using the T(X-sigma) scaling relation is in good agreement with the measured galaxy velocities. Our photometric redshift analysis suggests that the weak lensing signal observed to the south of the cluster and previously attributed to a ""dark clump"" is produced by background sources, possibly distributed as a filamentary structure.

The qWR star HD 45166 - II. Fundamental stellar parameters and evidence of a latitude-dependent wind

Context. The enigmatic object HD 45166 is a qWR star in a binary system with an orbital period of 1.596 day, and presents a rich emission-line spectrum in addition to absorption lines from the companion star (B7 V). As the system inclination is very small (i = 0.77 degrees +/- 0.09 degrees), HD 45166 is an ideal laboratory for wind-structure studies. Aims. The goal of the present paper is to determine the fundamental stellar and wind parameters of the qWR star. Methods. A radiative transfer model for the wind and photosphere of the qWR star was calculated using the non-LTE code CMFGEN. The wind asymmetry was also analyzed using a recently-developed version of CMFGEN to compute the emerging spectrum in two-dimensional geometry. The temporal-variance spectrum (TVS) was calculated to study the line-profile variations. Results. Abundances and stellar and wind parameters of the qWR star were obtained. The qWR star has an effective temperature of T(eff) = 50 000 +/- 2000 K, a luminosity of log(L/L(circle dot)) = 3.75 +/- 0.08, and a corresponding photospheric radius of R(phot) = 1.00 R(circle dot). The star is helium-rich (N(H)/N(He) = 2.0), while the CNO abundances are anomalous when compared either to solar values, to planetary nebulae, or to WR stars. The mass-loss rate is. M = 2.2 x 10(-7) M(circle dot) yr(-1), and the wind terminal velocity is v(infinity) = 425 km s(-1). The comparison between the observed line profiles and models computed under different latitude-dependent wind densities strongly suggests the presence of an oblate wind density enhancement, with a density contrast of at least 8: 1 from equator to pole. If a high velocity polar wind is present (similar to 1200 km s(-1)), the minimum density contrast is reduced to 4:1. Conclusions. The wind parameters determined are unusual when compared to O-type stars or to typical WR stars. While for WR stars v(infinity)/v(esc) > 1.5, in the case of HD 45166 it is much smaller (v(infinity)/v(esc) = 0.32). In addition, the efficiency of momentum transfer is eta = 0.74, which is at least 4 times smaller than in a typical WR. We find evidence for the presence of a wind compression zone, since the equatorial wind density is significantly higher than the polar wind. The TVS supports the presence of such a latitude-dependent wind and a variable absorption/scattering gas near the equator.

A NEW DIAGNOSTIC OF THE RADIAL DENSITY STRUCTURE OF Be DISKS

We analyze the intrinsic polarization of two classical Be stars in the process of losing their circumstellar disks via a Be to normal B star transition originally reported by Wisniewski et al. During each of five polarimetric outbursts which interrupt these disk-loss events, we find that the ratio of the polarization across the Balmer jump (BJ+/BJ-) versus the V-band polarization traces a distinct loop structure as a function of time. Since the polarization change across the Balmer jump is a tracer of the innermost disk density whereas the V-band polarization is a tracer of the total scattering mass of the disk, we suggest that such correlated loop structures in Balmer jump-V-band polarization diagrams (BJV diagrams) provide a unique diagnostic of the radial distribution of mass within Be disks. We use the three-dimensional Monte Carlo radiation transfer code HDUST to reproduce the observed clockwise loops simply by turning ""on/off"" the mass decretion from the disk. We speculate that counterclockwise loop structures we observe in BJV diagrams might be caused by the mass decretion rate changing between subsequent ""on/off"" sequences. Applying this new diagnostic to a larger sample of Be disk systems will provide insight into the time-dependent nature of each system's stellar decretion rate.

Spiral-like structure at the centre of nearby clusters of galaxies

Context. X-ray data analysis have found that fairly complex structures at cluster centres are more common than expected. Many of these structures have similar morphologies, which exhibit spiral-like substructure. Aims. It is not yet well known how these structures are formed or maintained. Understanding the origin of these spiral-like features at the centre of some clusters is the major motivation behind this work. Methods. We analyse deep Chandra observations of 15 nearby galaxy clusters ( 0.01 < z < 0.06), and use X-ray temperature and substructure maps to detect small features at the cores of the clusters. Results. We detect spiral-like features at the centre of 7 clusters: A85, A426, A496, Hydra A cluster, Centaurus, Ophiuchus, and A4059. These patterns are similar to those found in numerical hydrodynamic simulations of cluster mergers with non-zero impact parameter. In some clusters of our sample, a strong radio source also occupies the inner region of the cluster, which indicates a possible connection between the two. Our investigation implies that these spiral-like structures may be caused by off-axis minor mergers. Since these features occur in regions of high density, they may confine radio emission from the central galaxy producing, in some cases, unusual radio morphology.

Cyclic variability of the circumstellar disk of the Be star zeta Tauri II. Testing the 2D global disk oscillation model

Context. About 2/3 of the Be stars present the so-called V/R variations, a phenomenon characterized by the quasi-cyclic variation in the ratio between the violet and red emission peaks of the HI emission lines. These variations are generally explained by global oscillations in the circumstellar disk forming a one-armed spiral density pattern that precesses around the star with a period of a few years. Aims. This paper presents self-consistent models of polarimetric, photometric, spectrophotometric, and interferometric observations of the classical Be star zeta Tauri. The primary goal is to conduct a critical quantitative test of the global oscillation scenario. Methods. Detailed three-dimensional, NLTE radiative transfer calculations were carried out using the radiative transfer code HDUST. The most up-to-date research on Be stars was used as input for the code in order to include a physically realistic description for the central star and the circumstellar disk. The model adopts a rotationally deformed, gravity darkened central star, surrounded by a disk whose unperturbed state is given by a steady-state viscous decretion disk model. It is further assumed that this disk is in vertical hydrostatic equilibrium. Results. By adopting a viscous decretion disk model for zeta Tauri and a rigorous solution of the radiative transfer, a very good fit of the time-average properties of the disk was obtained. This provides strong theoretical evidence that the viscous decretion disk model is the mechanism responsible for disk formation. The global oscillation model successfully fitted spatially resolved VLTI/AMBER observations and the temporal V/R variations in the H alpha and Br gamma lines. This result convincingly demonstrates that the oscillation pattern in the disk is a one-armed spiral. Possible model shortcomings, as well as suggestions for future improvements, are also discussed.

Group I introns and associated homing endonuclease genes reveals a clinal structure for Porphyra spiralis var. amplifolia (Bangiales, Rhodophyta) along the Eastern coast of South America

Background: Group I introns are found in the nuclear small subunit ribosomal RNA gene (SSU rDNA) of some species of the genus Porphyra (Bangiales, Rhodophyta). Size polymorphisms in group I introns has been interpreted as the result of the degeneration of homing endonuclease genes (HEG) inserted in peripheral loops of intron paired elements. In this study, intron size polymorphisms were characterized for different Porphyra spiralis var. amplifolia (PSA) populations on the Southern Brazilian coast, and were used to infer genetic relationships and genetic structure of these PSA populations, in addition to cox2-3 and rbcL-S regions. Introns of different sizes were tested qualitatively for in vitro self-splicing. Results: Five intron size polymorphisms within 17 haplotypes were obtained from 80 individuals representing eight localities along the distribution of PSA in the Eastern coast of South America. In order to infer genetic structure and genetic relationships of PSA, these polymorphisms and haplotypes were used as markers for pairwise Fst analyses, Mantel's test and median joining network. The five cox2-3 haplotypes and the unique rbcL-S haplotype were used as markers for summary statistics, neutrality tests Tajima's D and Fu's Fs and for median joining network analyses. An event of demographic expansion from a population with low effective number, followed by a pattern of isolation by distance was obtained for PSA populations with the three analyses. In vitro experiments have shown that introns of different lengths were able to self-splice from pre-RNA transcripts. Conclusion: The findings indicated that degenerated HEGs are reminiscent of the presence of a full-length and functional HEG, once fixed for PSA populations. The cline of HEG degeneration determined the pattern of isolation by distance. Analyses with the other markers indicated an event of demographic expansion from a population with low effective number. The different degrees of degeneration of the HEG do not refrain intron self-splicing. To our knowledge, this was the first study to address intraspecific evolutionary history of a nuclear group I intron; to use nuclear, mitochondrial and chloroplast DNA for population level analyses of Porphyra; and intron size polymorphism as a marker for population genetics.

The Missing Part of Seed Dispersal Networks: Structure and Robustness of Bat-Fruit Interactions

Mutualistic networks are crucial to the maintenance of ecosystem services. Unfortunately, what we know about seed dispersal networks is based only on bird-fruit interactions. Therefore, we aimed at filling part of this gap by investigating bat-fruit networks. It is known from population studies that: (i) some bat species depend more on fruits than others, and (ii) that some specialized frugivorous bats prefer particular plant genera. We tested whether those preferences affected the structure and robustness of the whole network and the functional roles of species. Nine bat-fruit datasets from the literature were analyzed and all networks showed lower complementary specialization (H(2)' = 0.3760.10, mean 6 SD) and similar nestedness (NODF = 0.5660.12) than pollination networks. All networks were modular (M=0.32 +/- 0.07), and had on average four cohesive subgroups (modules) of tightly connected bats and plants. The composition of those modules followed the genus-genus associations observed at population level (Artibeus-Ficus, Carollia-Piper, and Sturnira-Solanum), although a few of those plant genera were dispersed also by other bats. Bat-fruit networks showed high robustness to simulated cumulative removals of both bats (R = 0.55 +/- 0.10) and plants (R = 0.68 +/- 0.09). Primary frugivores interacted with a larger proportion of the plants available and also occupied more central positions; furthermore, their extinction caused larger changes in network structure. We conclude that bat-fruit networks are highly cohesive and robust mutualistic systems, in which redundancy is high within modules, although modules are complementary to each other. Dietary specialization seems to be an important structuring factor that affects the topology, the guild structure and functional roles in bat-fruit networks.

Inferences of evolutionary and ecological events that influenced the population structure of Plebeia remota, a stingless bee from Brazil

The present study characterised the population genetic structure of Plebeia remota through mitochondrial DNA (mtDNA) analysis and evaluated evolutionary and ecological processes that may have contributed to the species current genetic scenario. Seventy feral nests were sampled representing four geographic regions (Cunha, Curitiba, Prudentopolis, and Blumenau). Fifteen composite mtDNA haplotypes were determined and a high genetic structure was detected among all populations. The current population structure may be a result of queen philopatry and vegetation shifts caused by palaeoclimatic changes and uplift of Brazilian coastal ranges. Finally, this study strongly suggests a revision of the taxonomic status of P. remota from the Prudentopolis region.

Phylogeography of the common vampire bat (Desmodus rotundus): Marked population structure, Neotropical Pleistocene vicariance and incongruence between nuclear and mtDNA markers

Background: The common vampire bat Desmodus rotundus is an excellent model organism for studying ecological vicariance in the Neotropics due to its broad geographic range and its preference for forested areas as roosting sites. With the objective of testing for Pleistocene ecological vicariance, we sequenced a mitocondrial DNA (mtDNA) marker and two nuclear markers (RAG2 and DRB) to try to understand how Pleistocene glaciations affected the distribution of intraspecific lineages in this bat. Results: Five reciprocally monophyletic clades were evident in the mitochondrial gene tree, and in most cases with high bootstrap support: Central America (CA), Amazon and Cerrado (AMC), Pantanal (PAN), Northern Atlantic Forest (NAF) and Southern Atlantic Forest (SAF). The Atlantic forest clades formed a monophyletic clade with high bootstrap support, creating an east/west division for this species in South America. On the one hand, all coalescent and non-coalescent estimates point to a Pleistocene time of divergence between the clades. On the other hand, the nuclear markers showed extensive sharing of haplotypes between distant localities, a result compatible with male-biased gene flow. In order to test if the disparity between the mitochondrial and nuclear markers was due to the difference in mutation rate and effective size, we performed a coalescent simulation to examine the feasibility that, given the time of separation between the observed lineages, even with a gene flow rate close to zero, there would not be reciprocal monophyly for a neutral nuclear marker. We used the observed values of theta and an estimated mutation rate for the nuclear marker gene to perform 1000 iterations of the simulation. The results of this simulation were inconclusive: the number of iterations with and without reciprocal monophyly of one or more clades are similar. Conclusions: We therefore conclude that the pattern exhibited by the common vampire bat, with marked geographical structure for a mitochondrial marker and no phylogeographic structure for nuclear markers is compatible with a historical scenario of complete isolation of refuge-like populations during the Pleistocene. The results on demographic history on this species is compatible with the Carnaval-Moritz model of Pleistocene vicariance, with demographic expansions in the southern Atlantic forest.