Variations on a theme: diversification of cuticular hydrocarbons in a clade of cactophilic Drosophila

Background: We characterized variation and chemical composition of epicuticular hydrocarbons (CHCs) in the seven species of the Drosophila buzzatii cluster with gas chromatography/mass spectrometry. Despite the critical role of CHCs in providing resistance to desiccation and involvement in communication, such as courtship behavior, mating, and aggregation, few studies have investigated how CHC profiles evolve within and between species in a phylogenetic context. We analyzed quantitative differences in CHC profiles in populations of the D. buzzatii species cluster in order to assess the concordance of CHC differentiation with species divergence. Results: Thirty-six CHC components were scored in single fly extracts with carbon chain lengths ranging from C(29) to C(39), including methyl-branched alkanes, n alkenes, and alkadienes. Multivariate analysis of variance revealed that CHC amounts were significantly different among all species and canonical discriminant function (CDF) analysis resolved all species into distinct, non-overlapping groups. Significant intraspecific variation was found in different populations of D. serido suggesting that this taxon is comprised of at least two species. We summarized CHC variation using CDF analysis and mapped the first five CHC canonical variates (CVs) onto an independently derived period (per) gene + chromosome inversion + mtDNA COI gene for each sex. We found that the COI sequences were not phylogenetically informative due to introgression between some species, so only per + inversion data were used. Positive phylogenetic signal was observed mainly for CV1 when parsimony methods and the test for serial independence (TFSI) were used. These results changed when no outgroup species were included in the analysis and phylogenetic signal was then observed for female CV3 and/or CV4 and male CV4 and CV5. Finally, removal of divergent populations of D. serido significantly increased the amount of phylogenetic signal as up to four out of five CVs then displayed positive phylogenetic signal. Conclusions: CHCs were conserved among species while quantitative differences in CHC profiles between populations and species were statistically significant. Most CHCs were species-, population-, and sex-specific. Mapping CHCs onto an independently derived phylogeny revealed that a significant portion of CHC variation was explained by species' systematic affinities indicating phylogenetic conservatism in the evolution of these hydrocarbon arrays, presumptive waterproofing compounds and courtship signals as in many other drosophilid species.

Functional characterization of the sciarid BhC4-1 core promoter in transgenic Drosophila

Background: Core promoters are cis-regulatory modules to which bind the basal transcriptional machinery and which participate in the regulation of transcription initiation. Although core promoters have not been extensively investigated through functional assays in a chromosomal context, the available data suggested that the response of a given core promoter might vary depending on the promoter context. Previous studies suggest that a (-57/+40) fragment constitutes the core promoter of the BhC4-1 gene which is located in DNA puff C4 of the sciarid fly Bradysia hygida. Here we tested this (-57/+40) fragment in distinct regulatory contexts in order to verify if promoter context affects its core promoter activity. Results: Consistent with the activity of a core promoter, we showed that in the absence of upstream regulatory sequences the (-57/+40) fragment drives low levels of reporter gene mRNA expression throughout development in transgenic Drosophila. By assaying the (-57/+40) fragment in two distinct regulatory contexts, either downstream of the previously characterized Fbp1 enhancer or downstream of the UAS element, we showed that the BhC4-1 core promoter drives regulated transcription in both the germline and in various tissues throughout development. Furthermore, the use of the BhC4-1 core promoter in a UAS construct significantly reduced salivary gland ectopic expression in third instar larvae, which was previously described to occur in the context of the GAL4/UAS system. Conclusions: Our results from functional analysis in transgenic Drosophila show that the BhC4-1 core promoter drives gene expression regardless of the promoter context that was assayed. New insights into the functioning of the GAL4/UAS system in Drosophila were obtained, indicating that the presence of the SV40 sequence in the 3' UTR of a UAS construct does not preclude expression in the germline. Furthermore, our analysis indicated that ectopic salivary gland expression in the GAL4/UAS system does not depend only on sequences present in the GAL4 construct, but can also be affected by the core promoter sequences in the UAS construct. In this context, we propose that the sciarid BhC4-1 core promoter constitutes a valuable core promoter which can be employed in functional assays in insects.

Deep sequencing of New World screw-worm transcripts to discover genes involved in insecticide resistance

Background: The New World screw-worm (NWS), Cochliomyia hominivorax, is one of the most important myiasis-causing flies, causing severe losses to the livestock industry. In its current geographical distribution, this species has been controlled by the application of insecticides, mainly organophosphate (OP) compounds, but a number of lineages have been identified that are resistant to such chemicals. Despite its economic importance, only limited genetic information is available for the NWS. Here, as a part of an effort to characterize the C. hominivorax genome and identify putative genes involved in insecticide resistance, we sampled its transcriptome by deep sequencing of polyadenylated transcripts using the 454 sequencing technology. Results: Deep sequencing on the 454 platform of three normalized libraries (larval, adult male and adult female) generated a total of 548,940 reads. Eighteen candidate genes coding for three metabolic detoxification enzyme families, cytochrome P450 monooxygenases, glutathione S transferases and carboxyl/cholinesterases were selected and gene expression levels were measured using quantitative real-time polymerase chain reaction (qRT-PCR). Of the investigated candidates, only one gene was expressed differently between control and resistant larvae with, at least, a 10-fold down-regulation in the resistant larvae. The presence of mutations in the acetylcholinesterase (target site) and carboxylesterase E3 genes was investigated and all of the resistant flies presented E3 mutations previously associated with insecticide resistance. Conclusions: Here, we provided the largest database of NWS expressed sequence tags that is an important resource, not only for further studies on the molecular basis of the OP resistance in NWS fly, but also for functional and comparative studies among Calliphoridae flies. Among our candidates, only one gene was found differentially expressed in resistant individuals, and its role on insecticide resistance should be further investigated. Furthermore, the absence of mutations in the OP target site and the high frequency of mutant carboxylesterase E3 indicate that metabolic resistance mechanisms have evolved predominantly in this species.

Gene Expression Noise in Spatial Patterning: hunchback Promoter Structure Affects Noise Amplitude and Distribution in Drosophila Segmentation

Positional information in developing embryos is specified by spatial gradients of transcriptional regulators. One of the classic systems for studying this is the activation of the hunchback (hb) gene in early fruit fly (Drosophila) segmentation by the maternally-derived gradient of the Bicoid (Bcd) protein. Gene regulation is subject to intrinsic noise which can produce variable expression. This variability must be constrained in the highly reproducible and coordinated events of development. We identify means by which noise is controlled during gene expression by characterizing the dependence of hb mRNA and protein output noise on hb promoter structure and transcriptional dynamics. We use a stochastic model of the hb promoter in which the number and strength of Bcd and Hb (self-regulatory) binding sites can be varied. Model parameters are fit to data from WT embryos, the self-regulation mutant hb(14F), and lacZ reporter constructs using different portions of the hb promoter. We have corroborated model noise predictions experimentally. The results indicate that WT (self-regulatory) Hb output noise is predominantly dependent on the transcription and translation dynamics of its own expression, rather than on Bcd fluctuations. The constructs and mutant, which lack self-regulation, indicate that the multiple Bcd binding sites in the hb promoter (and their strengths) also play a role in buffering noise. The model is robust to the variation in Bcd binding site number across a number of fly species. This study identifies particular ways in which promoter structure and regulatory dynamics reduce hb output noise. Insofar as many of these are common features of genes (e. g. multiple regulatory sites, cooperativity, self-feedback), the current results contribute to the general understanding of the reproducibility and determinacy of spatial patterning in early development.

Role of the gp85/Trans-Sialidases in Trypanosoma cruzi Tissue Tropism: Preferential Binding of a Conserved Peptide Motif to the Vasculature In Vivo

Background: Transmitted by blood-sucking insects, the unicellular parasite Trypanosoma cruzi is the causative agent of Chagas' disease, a malady manifested in a variety of symptoms from heart disease to digestive and urinary tract dysfunctions. The reasons for such organ preference have been a matter of great interest in the field, particularly because the parasite can invade nearly every cell line and it can be found in most tissues following an infection. Among the molecular factors that contribute to virulence is a large multigene family of proteins known as gp85/trans-sialidase, which participates in cell attachment and invasion. But whether these proteins also contribute to tissue homing had not yet been investigated. Here, a combination of endothelial cell immortalization and phage display techniques has been used to investigate the role of gp85/trans-sialidase in binding to the vasculature. Methods: Bacteriophage expressing an important peptide motif (denominated FLY) common to all gp85/trans-sialidase proteins was used as a surrogate to investigate the interaction of this motif with the endothelium compartment. For that purpose phage particles were incubated with endothelial cells obtained from different organs or injected into mice intravenously and the number of phage particles bound to cells or tissues was determined. Binding of phages to intermediate filament proteins has also been studied. Findings and Conclusions: Our data indicate that FLY interacts with the endothelium in an organ-dependent manner with significantly higher avidity for the heart vasculature. Phage display results also show that FLY interaction with intermediate filament proteins is not limited to cytokeratin 18 (CK18), which may explain the wide variety of cells infected by the parasite. This is the first time that members of the intermediate filaments in general, constituted by a large group of ubiquitously expressed proteins, have been implicated in T. cruzi cell invasion and tissue homing.

CHEMICAL AND PHYSICO-CHEMICAL CHARACTERIZATION OF LIGNINS OBTAINED FROM ETHANOL-WATER FRACTIONATION OF BAGASSE

Organosolv lignins can replace petroleum chemicals such as phenol either partially or totally in various applications. Eight lignins, seven of which corresponded to the ethanol-water fractionation of bagasse and the other to a reference lignin (Alcell (R)) were analyzed with the aim to evaluate their chemical and physicochemical characteristics. The purity of the lignin fractions was determined by high pressure liquid chromatography (HPLC) and by ash content. Fourier Transform-Infrared Spectroscopy (FTIR) techniques and differential UV spectroscopy were applied to identify the chemical groups in the lignin samples. The molecular weight distribution was determined by size exclusion chromatography (HPSEC). Thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) techniques were used to determine the mass loss due to the high temperature treatment. The lignins studied showed the presence of p-hydroxyphenyl (H unit) and a greater proportion of guaiacyl (G unit) moieties, lower purity, similar or greater amount of phenolic hydroxyl groups, and higher degradation temperatures, than the Alcell (R) lignin.

A New Generation of X Ray Irradiators for Insect Sterilization

Recent fears of terrorism have provoked an increase in delays and denials of transboundary shipments of radioisotopes. This represents a serious constraint to sterile insect technique (SIT) programs around the world as they rely on the use of ionizing radiation from radioisotopes for insect sterilization. To validate a novel X ray irradiator, a series of studies on Ceratitis capitata (Wiedemann) and Anastrepha fraterculus (Wiedemann) (Diptera: Tephritidae) were carried out, comparing the relative biological effectiveness (RBE) between X rays and traditional gamma radiation from (60)Co. Male C. capitata pupae and pupae of both sexes of A. fraterculus, both 24 - 48 h before adult emergence, were irradiated with doses ranging from 15 to 120 Gy and 10-70 Gy, respectively. Estimated mean doses of 91.2 Gy of X and 124.9 Gy of gamma radiation induced 99% sterility in C. capitata males, Irradiated A. fraterculus were 99% sterile at approximate to 40-60 Gy for both radiation treatments. Standard quality control parameters and mating indices were not significantly affected by the two types of radiation. The RBE did not differ significantly between the tested X and gamma radiation, and X rays are as biologically effective for SIT purposes as gamma rays are. This work confirms the suitability of this new generation of X ray irradiators for pest control programs that integrate the SIT.

Intraspecific queen parasitism in a highly eusocial bee

Insect societies are well-known for their advanced cooperation, but their colonies are also vulnerable to reproductive parasitism. Here, we present a novel example of an intra-specific social parasitism in a highly eusocial bee, the stingless bee Melipona scutellaris. In particular, we provide genetic evidence which shows that, upon loss of the mother queen, many colonies are invaded by unrelated queens that fly in from unrelated hives nearby. The reasons for the occurrence of this surprising form of social parasitism may be linked to the fact that unlike honeybees, Melipona bees produce new queens in great excess of colony needs, and that this exerts much greater selection on queens to seek alternative reproductive options, such as by taking over other nests. Overall, our results are the first to demonstrate that queens in highly eusocial bees can found colonies not only via supersedure or swarming, but also by infiltrating and taking over other unrelated nests.

Lightweight composite concrete produced with water treatment sludge and sawdust: Thermal properties and potential application

The main objective of this study was to evaluate the potential application of a lightweight concrete produced with lightweight coarse aggregate made of the water treatment sludge and sawdust (lightweight composite), by determining the thermal properties and possible environmental impact of future residue of this concrete. Two types of concrete were prepared: concrete produced with the lightweight composite dosed with cement/sand/composite/water in a mass ratio of 1:2.5:0.67:0.6 and conventional concrete dosed with cement/sand/crushed stone/water in a mass ratio of 1:4.8:5.8:0.8. The thermal properties were determined by the hot wire parallel technique. The possible environmental impact was measured using the procedures and guidelines of the Brazilian Association of Technical Standards - ABNT. The concrete produced with the lightweight composite presented a 23% lower thermal conductivity than the conventional concrete. The concrete produced with the lightweight composite presented a set of thermal properties suitable for the application of this concrete in non-structural sealing elements. The concentration of aluminum in the solubilized extract of the concrete produced with the lightweight composite was much lower than the concentration of aluminum in the water treatment sludge, confirming the possible reduction of environmental impact of this composite for use in concrete. (C) 2010 Elsevier Ltd. All rights reserved.

Laboratory measurements of biomarkers and individual performances in Chironomus xanthus to evaluate pesticide contamination of sediments in a river of southeastern Brazil

This study aimed at evaluating biomarkers, individual and population responses in the native Chironomus xanthus to assess the toxicity of pesticide-contaminated sediments from the Monjolinho River (Southeast Brazil). We measured cholinesterase (ChE) and glutathione S-transferase activities (GST), as biomarkers and survival, individual growth and adult emergence, as individual performances. There was no response of the ChE activity and a tendency to decreased GST activity in contaminated sites, but this was generally not statistically significant. Therefore, there was no association of the biomarker responses with exposure to sediment containing pesticides. In contrast, ash free dry mass was significantly increased and male emergence was decreased in C. xanthus exposed to the same sediments. In conclusion, the selected biomarkers were not sensitive and specific enough to detect and anticipate effects of pesticide contamination at the levels measured in the study area. Nevertheless, individual performances alterations pointed to potential pollution problems and possible ecological consequences. (C) 2010 Elsevier Inc. All rights reserved.

Effects of temperature at different organic loading levels on the performance of a fluidized-bed anaerobic sequencing batch bioreactor

An investigation was performed on the effect of temperature and organic load on the stability and efficiency of a 1.8-L fluidized-bed anaerobic sequencing batch reactor (ASBR), containing granulated biomass. Assays were carried out employing superficial up How velocity of 7 m/h, total cycle length of 6 h and synthetic wastewater volume of 1.3 L per cycle. The fluidized-bed ASH was operated at 15, 20, 25 and 30 degrees C with influent organic matter concentrations of 500 and 1000 mgCOD/L The system showed stability under all conditions and presented filtered samples removal efficiency ranging from 79 to 86%. A first-order kinetic model could be fitted to the experimental values of the organic matter concentration profiles. The specific kinetic parameter values of this model ranged from 0.0435 to 0.2360 L/(gTVS h) at the implemented operation conditions. in addition, from the slope of an Arrhenius plot, the activation energy values were calculated to be 16,729 and 12,673 cal/mol for operation with 500 and 1000 mgCOD/L, respectively. These results show that treatment of synthetic wastewater. with concentration of 500 mgCOD/L, was more sensitive to temperature variations than treatment of the same residue with concentration of 1000 mgCOD/L. Comparing the activation energy value for operation at 500 mgCOD/L with the value obtained by Agibert et al. (S.A. Agibert, M.B. Moreira, S.M. Ratusznei, J.A.D. Rodrigues, M. Zaiat, E. Foresti. Influence of temperature on performance of an ASBBR with circulation applied to treatment of low-strength wastewater. journal of Applied Biochemistry and Biotechnology, 136 (2007) 193-206) in an ASBBR treating the same wastewater at the same concentration, the value obtained in the fluidized-bed ASBR showed to be superior, indicating that treatment of synthetic wastewater in a reactor containing granulated biomass was more sensitive to temperature variations than the treatment using immobilized biomass. (c) 2008 Elsevier B.V. All rights reserved.

Sources of Variations When Comparing Concrete Carbonation Results

Carbonation is one of the main concerns for concrete service life in tropical countries. The mechanism and materials that produce it have been widely studied as well as natural and accelerated methods to report and analyze it. In spite of reported investigations, there is a need for information that could allow an adequate interpretation of the results of the standardization process. This lack of information can produce variations not only in the interpretation but also in the predictions of service life. The purpose of this paper is to analyze and discuss variables that could be sources of error, especially when performing accelerated tests. As a result, a methodologies to minimize variations when interpreting and comparing results is proposed, such as specimen geometry and preconditioning, spacing, relative humidity, and CO(2) concentration.

Froth flotation of coal tailings

The processes involved in coal preparation produce harmful effects for the environment, mainly by the quantity and nature Of wastes that are generated, These tailings generally called ""pyritic"", even though not presenting a high pyrite content, are an acid material harmful to the environment. These tailings are usually disposed of in tailings dams. The tailings, studied in this work come from a process involving dense-medium cyclones and spirals. They have an ash content of 56% and a calorific value of 5,800 BTU/Lb, the sulfur content is 1.2%. In terms of size, the material is considered as ultrafine, as 63% of it is less than 0,014 mm. The coal matter content of these tailings is easily recovered by froth flotation, as evidenced in this work. It possible to recover 74% of the coal matter and to obtain a product with 7.3% ash and calorific value of 14,225 BTU/lb in dry basis.

CALCINED SLUDGE SINTERING EVALUATION BY HEATING MICROSCOPY THERMAL ANALYSIS

Aiming the use of the sewage sludge produced in one of the largest Brazilian wastewater treatment stations as a raw material for the ceramic industry, the sintering process of the ashes produced from its calcination was evaluated by heating microscopy thermal analysis (HMTA). From the microprocessed images, a method was developed to obtain HMTA dimensional change curves as a function of temperature, equivalent to those usually obtained from dilatometers or by thermomechanical analysis (TMA). The final product after sintering at 1050 degrees C, characterized by X-ray fluorescence spectrometry, scanning electron microscopy and X-ray dispersive energy, indicates the presence of a vitreous phase containing phosphorus, which explains the good sintering properties of the studied calcined sludge, as shown from its HMTA dimensional change curve.

Biogenic modified silica as a sorbent of cadmium ions: Preparation and characterization

This work describes the preparation and characterization of biogenic modified silica from rice hull ash and its use as a sorbent of cadmium ions. Thus, an agro-industrial residue has been used to produce a new adsorbent product which is able to remove toxic elements. Mesoporous biogenic silica was obtained by alkaline extraction of sodium silicate by hydrolysis with the sol-gel process, and it was modified with salen using 1,2-dichloroethane as a spacer. The surface area of the silica was measured by nitrogen adsorption/desorption analysis. Surface modification was measured by Fourier transform infrared spectroscopy. The degree of functionalization was obtained by elemental analysis. This work showed that biogenic modified silica can be produced in aqueous media from rice hull ash using a simple method, providing an alternative method for adsorbent preparation. Thermogravimetric analysis showed that the salen-modified silica is stable up to 209 C. The modified silica displays appropriate structural characteristics for an adsorbent. The cylindrical pores, open at both ends, allow free diffusion of cadmium ions to the adsorption sites on the silica surface. The surface modification increases cadmium adsorption on the silica surface 100-fold. The salen-modified silica showed specific adsorption for Cd2+ of 44.52 mg/g SiO2 at cadmium concentration of 100 mg/l.