Reverse supply chain practices for construction and demolition waste in the Brazilian Amazon: a multi-stakeholder view.

Construction and demolition waste (CDW) is an environmental problem that affects all regions of the world. Particularly in the Brazilian Amazon Forest region, the volume of CDW generated almost doubled between 2007 and 2019. Indeed, despite Brazil having environmental regulations for waste management, these have been insufficient to solve the environmental problem because there is no CDW reverse supply chain (RSC) properly developed in the Amazon region. Previous studies have proposed a conceptual model of a CDW RSC but have hitherto failed to apply them against real world practice. This paper, therefore, attempts to test existing conceptual models that describe a CDW RSC against real industry practice prior to developing an applied model of a CDW RSC for the Brazilian Amazon. To modify the conceptual model for CDW RSC, qualitative data through 15 semi-structured interviews with five different types of stakeholders of the Amazonian CDW RSC were collected and analyzed using qualitative content analysis methods using NVivo software. The proposed applied model includes present and future reverse logistics (RL) practices, and strategies and tasks necessary for the implementation of a CDW RSC in the city of Belém of Pará, in the Brazilian Amazon. Findings reveal that several overlooked problems, particularly the limitations of the existing legal framework in Brazil, are not enough to promote a robust CDW RSC. This is perhaps the first study to examine CDW RSC in the Amazonian rainforest. Arguments provided in this study highlight the necessity for an Amazonian CDW RSC that must be promoted and regulated by the government. This can be addressed by the utilizing public-private partnership (PPP) for developing a CDW RSC.

Antifungal activity of poly(ε-caprolactone) nanoparticles incorporated with Eucalyptus essential oils against Hemileia vastatrix.

Coffee (Coffea L.) is one of the main crops produced globally. Its contamination by the fungus Hemileia vastatrix Berkeley and Broome has been economically detrimental for producers. The objective of this work was to extract and characterize the essential oils from Eucalyptus citriodora Hook, Eucalyptus camaldulensis Dehn and Eucalyptus grandis Hill ex Maiden, produce and characterize nanoparticles containing these essential oils and evaluate the in vivo and in vitro antifungal activity of free and nanoencapsulated essential oils. The principal constituent of the essential oil from E. citriodora was citronellal; that from E. grandis was α-pinene; and that from E. camaldulensis was 1,8-cineol. The in vitro antifungal activity against the fungus H. vastatrix was 100% at a concentration of 1000 µl l-1 for all the oils and nanoparticles containing these natural products. The sizes of the nanoparticles produced with the essential oils from E. citriodora, E. camaldulensis and E. grandis were 402·13 nm, 275·33 nm and 328·5 nm, respectively, with surface charges of -11·8 mV, -9·24 mV and - 6·76 mV, respectively. Fourier transform infrared analyses proved that the encapsulation of essential oils occurred in the polymeric matrix of poly(ε-caprolactone). The incorporation of essential oils into biodegradable poly(ε-caprolactone) nanoparticles increased their efficiency as biofungicides in the fight against coffee rust, decreasing the severity of the disease by up to 90·75% after treatment with the nanoparticles containing the essential oil from E. grandis.

Absorption characteristics of large acoustic metasurfaces.

Metasurfaces formed of arrays of subwavelength resonators are often tuned to 'critically couple' with incident radiation, so that at resonance dissipative and radiative damping are balanced and absorption is maximized. Such design criteria are typically derived assuming an infinite metasurface, whereas the absorption characteristics of finite metasurfaces, even very large ones, can be markedly different in certain frequency intervals. This is due to the excitation of surface waves, intrinsic to resonant metasurfaces and especially meta-resonances, namely collective resonances where the surface waves form standing-wave patterns over the planar metasurface domain. We illustrate this issue using a detailed model of a Helmholtz-type acoustic metasurface formed of cavity-neck pairs embedded into a rigid substrate, with geometric and dissipation effects included from first principles (R. Brandão and O. Schnitzer, Wave Motion, 97, 102583, 2020). This article is part of the theme issue 'Wave generation and transmission in multi-scale complex media and structured metamaterials (part 1)'.

Synchronization performance affects gait variability measures during cued walking.

BACKGROUND: Incorporating variability within gait rehabilitation offers a promising approach to restore functional capacity. However, it's success requires adequate synchronization, a parameter that lacks report in most of the literature regarding cued gait training. RESEARCH QUESTION: How changes to synchronization performance during fractal-like and isochronous cueing impacts gait variability measures? METHODS: We asked twelve young male participants to walk in synchronization to two different temporally structure cueing (isochronous [ISO] and fractal [FRC]). We have also manipulated the cueing's tempo by increasing and decreasing it by 5% to manipulate synchronization, resulting in six conditions (stimuli [ISO,FRC] x tempo [SLOW, NORMAL, FAST]). The normal condition was set from an uncued trial through the participant's self-paced stride time. Synchronization performance (ASYNC) and gait variability (fractal scaling and coefficient of variation) were calculated from stride time data ( -ISIs,CV-ISIs). Repeated measures analysis of variance or Aligned Rank Transform were conducted to determine significant differences between metronome tempo and stimuli for the dependent variables RESULTS: Our results showed a FAST tempo decreases synchronization performance (ASYNC) and leads to lower -ISIs, for both ISO and FRC stimuli. This indicates that when an individual exhibits poor synchronization during cued gait training, his/her gait variability patterns will not follow the temporal structure of the presented metronome. Specifically, if the individual poorly synchronizes to the cues, the gait patterns become more random, a condition typically observed in older adults and neurological patients, which runs contrary to the hypothesis when using fractal-like metronomes. SIGNIFICANCE: This study provides supporting evidence that measuring synchronization performance in cued training is fundamental for a proper clinical interpretation of its effects. This is particularly relevant for the recent and ongoing clinical research using fractal-like metronomes since the expected gait patterns are dependent on the synchronization performance. Randomized control trials must incorporate synchronization performance related measures.

Outcome measurement instruments in Rett syndrome: A systematic review.

OBJECTIVE: The aim of the study was to identify and characterize outcome measures for objective and subjective assessment in persons with Rett syndrome (RS). METHODS: A systematic review was conducted consulting the EBSCO, Cochrane, Web of Science, Scielo, MEDLINE and PsycINFO databases for published studies describing the use of patient-reported outcome measures (PROMs) and other outcome measures in persons with RS. Validation studies and observational studies were included. The PROMs were first described, and then the measurement properties were evaluated using predefined criteria according to the COnsensus-based Standards for the selection of health Measurement Instruments (COSMIN). The outcome measures were then grouped according to the International Classification of Functioning, Disability and Health (ICF) to establish a relationship between outcome measures and ICF domains. RESULTS: Twenty out of 2327 articles were appraised, and seventeen different outcome measures were identified and described. Ten outcome measures corresponded to evaluation questionnaires, while the remaining seven assessed functional outcomes: walking distance, physical activity level and ability to interact visually. A relation between these outcome measures that assess RS and the ICF allows understanding that most of the instruments (fifteen) include the assessment of activity limitations. CONCLUSIONS: The findings of this study seem to be promising for their use by clinicians and researchers, although they have methodological limitations. The accuracy and quality of these individual outcome measures should continue to be assessed in an attempt to gather a consensus on the best tools used in RS.

Antifungal and antiocratoxigenic potential of Alpinia speciosa and Cymbopogon flexuosus essential oils encapsulated in poly(lactic acid) nanofibres against Aspergillus fungi.

Essential oils encapsulated in a polymeric matrix can be used as an alternative method to control fungi and mycotoxins. The essential oils were extracted by hydrodistillation and characterized by gas chromatography. The nanofibres were produced from poly (acid lactic) (PLA) containing essential oils by the Solution Blow Spinning method. The antifungal and antimicotoxygenic properties were evaluated against Aspergillus ochraceus and Aspergillus westerdijkiae by the fumigation method. Terpinen-4-ol (20·23%), sabinene (20·18%), 1·8-cineole (16·69%) and γ-terpinene (11·03%) were the principal compounds present in the essential oil from Alpinia speciosa, whereas citral (97·67%) was dominant from Cymbopogon flexuosus. Microscopy images showed that the addition of essential oils caused an increase in the diameter of the nanofibres. The infrared spectroscopy results indicated the presence of essential oils in the PLA nanofibres. Differential scanning calorimetry curves also indicated the existence of interactions between the essential oils and polymeric macromolecules through their plasticizing action. The hydrophobic character of nanofibres was revealed by the contact angle technique. An antifungal effect was observed, the mycelial growths (3·25-100%) and the synthesis of ochratoxin A (25·94-100%) were inhibited by the presence of the nanofibres. The results suggest that bioactive nanofibres hold promise for application to control toxigenic fungi.

Bactericidal and antioxidant effects of essential oils from Satureja montana L., Myristica fragrans H. and Cymbopogon flexuosus.

The extraction and characterization of the essential oils (EO) from Satureja montana L., Myristica fragrans H. and Cymbopogon flexuosus and the determination of their antibacterial and antioxidant activities were achieved. The EO were identified by gas chromatography/mass spectrometry and quantified by gas chromatography using a flame ionization detector. The antibacterial potential against Escherichia coli and Staphylococcus aureus was evaluated by cell susceptibility assays and by scanning electron microscopy. The antioxidant activity was evaluated by the 2,2-diphenyl-1-picrylhydrazyl assay, by ß-carotene bleaching and by determining the reducing power. Borneol (36·18%), γ-terpineol (12·66%) and carvacrol (11·07%) were the principal components in the EO from S. montana, and sabinene (49·23%) and α-pinene (13·81%) were found in the EO from M. fragrans. Geranial (59·66%) and neral (38·98%) isomers were the only major components in the EO from C. flexuosus. The EO from S. montana was effective against E. coli, with minimum inhibitory and bactericidal concentrations (MIC and MBC) of 6·25 µl ml-1 , whereas bactericidal potential against both was observed for the EO from M. fragrans; MIC = 6·25 µl ml-1 for S. aureus and MBC = 12·5 µl ml-1 for E. coli. A significant protective role on lipid substrates in the ß-carotene bleaching assay was seen for the EO from S. montana and M. fragrans. Overall, such EO can be promising agents against pathogenic bacteria and for protecting biomolecules during oxidative stress.

Pathogenic neurofibromatosis type 1 (NF1) RNA splicing resolved by targeted RNAseq.

Neurofibromatosis type 1 (NF1) is caused by loss-of-function variants in the NF1 gene. Approximately 10% of these variants affect RNA splicing and are either missed by conventional DNA diagnostics or are misinterpreted by in silico splicing predictions. Therefore, a targeted RNAseq-based approach was designed to detect pathogenic RNA splicing and associated pathogenic DNA variants. For this method RNA was extracted from lymphocytes, followed by targeted RNAseq. Next, an in-house developed tool (QURNAs) was used to calculate the enrichment score (ERS) for each splicing event. This method was thoroughly tested using two different patient cohorts with known pathogenic splice-variants in NF1. In both cohorts all 56 normal reference transcript exon splice junctions, 24 previously described and 45 novel non-reference splicing events were detected. Additionally, all expected pathogenic splice-variants were detected. Eleven patients with NF1 symptoms were subsequently tested, three of which have a known NF1 DNA variant with a putative effect on RNA splicing. This effect could be confirmed for all 3. The other eight patients were previously without any molecular confirmation of their NF1-diagnosis. A deep-intronic pathogenic splice variant could now be identified for two of them (25%). These results suggest that targeted RNAseq can be successfully used to detect pathogenic RNA splicing variants in NF1.

Quality improvement and geographical indication of cachaça (Brazilian spirit) by using locally selected yeast strains.

AIMS: In order to improve the quality and to create a biological basis for obtainment of the protected denomination of origin (PDO), indigenous yeast were isolated and characterized for use in Salinas city (the Brazilian region of quality cachaça production). MATERIAL AND METHODS: Seven thousand and two hundred yeast colonies from 15 Salinas city distilleries were screened based on their fermentative behaviour and the physicochemical composition of cachaça. Molecular polymorphic analyses were performed to characterize these isolates. RESULTS: Two Saccharomyces cerevisiae strains (nos. 678 and 680) showed appropriate characteristics to use in the cachaça production: low levels of acetaldehyde and methanol, and high ethyl lactate/ethyl acetate ratio respectively. They also presented polymorphic characteristics more closely related between themselves even when compared to other strains from Salinas. CONCLUSIONS: The application of selected yeast to cachaça production can contribute for the improvement of the quality product as well as be used as a natural marker for PDO. SIGNIFICANCE AND IMPACT OF THE STUDY: This study suggests that the use of selected yeast strains could contribute to obtain a cachaça similar to those produced traditionally, while getting wide acceptation in the market, yet presenting more homogeneous organoleptic characteristics, and thus contributing to the PDO implementation.

Kinetics and regulation of lactose transport and metabolism in Kluyveromyces lactis JA6.

Kluyveromyces lactis strains are able to assimilate lactose. They have been used industrially to eliminate this sugar from cheese whey and in other industrial products. In this study, we investigated specific features and the kinetic parameters of the lactose transport system in K. lactis JA6. In lactose grown cells, lactose was transported by a system transport with a half-saturation constant (K s) of 1.49 ± 0.38 mM and a maximum velocity (V max) of 0.96 ± 0.12 mmol. (g dry weight)(-1) h(-1) for lactose. The transport system was constitutive and energy-dependent. Results obtained by different approaches showed that the lactose transport system was regulated by glucose at the transcriptional level and by glucose and other sugars at a post-translational level. In K. lactis JA6, galactose metabolization was under glucose control. These findings indicated that the regulation of lactose-galactose regulon in K. lactis was similar to the regulation of galactose regulon in Saccharomyces cerevisiae.

FMR1 low sub-genotype does not rescue BRCA1/2-mutated human embryos and does not explain primary ovarian insufficiency among BRCA1/2-carriers.

STUDY QUESTION: Can we confirm in our population whether FMRI low sub-genotypes are associated with BRCA1/2 mutations, as recently proposed? SUMMARY ANSWER: Our results indicate that the distribution of the FMR1 sub-genotypes in female BRCA1/2-mutation carriers is significantly different from what has been reported previously and resembles that of the control population. FMRI low sub-genotypes are not associated with BRCA1/2 mutations and this association is also absent among male mutation carriers. WHAT IS KNOWN ALREADY: Recently, BRCA1 mutations were reported to be associated with primary ovarian insufficiency (POI) in female carriers. In animal models, BRCA2-deficiency also results in impaired oogenesis. A recent study has reported that the POI in BRCA1/2-mutation carriers is most likely due to low FMR1 sub-genotype (CGG n < 26) and the authors also suggested that low sub-genotypes of the FMR1 gene might be important to rescue the BRCA1/2 embryos, which would otherwise be embryonically-lethal. STUDY DESIGN, SIZE, DURATION: This retrospective study was performed in October and November of 2012, using genetic material of 464 patients who underwent genetic screening in our centre in the past. PARTICIPANTS/MATERIALS, SETTING, METHODS: We tested the FMR1 sub-genotypes in 60 female and 29 males with either BRCA1 or BRCA2 mutations and 375 controls by PCR amplification and size fragment analysis. MAIN RESULTS: We did not find any evidence for an association of FMR1 low sub-genotypes and BRCA1/2 mutations. LIMITATIONS, REASONS FOR CAUTION: This association study assumes that the female BRCA1/2 population tested has POI. WIDER IMPLICATIONS OF THE FINDINGS: Low FMR1 sub-genotypes are not responsible for the presumed rescue of embryos with BRCA1/2 mutations. Furthermore, the molecular mechanism of the POI in BRCA1/2-female carriers is not likely to be associated with low FMR1 sub-genotype. STUDY FUNDING/COMPETING INTEREST(S): The Department of Clinical Genetics of the Maastricht University Medical Centre supported the study. The authors do not have any competing interests to declare.

Airborne olive pollen counts are not representative of exposure to the major olive allergen Ole e 1.

Pollen is routinely monitored, but it is unknown whether pollen counts represent allergen exposure. We therefore simultaneously determined olive pollen and Ole e 1 in ambient air in Córdoba, Spain, and Évora, Portugal, using Hirst-type traps for pollen and high-volume cascade impactors for allergen. Pollen from different days released 12-fold different amounts of Ole e 1 per pollen (both locations P < 0.001). Average allergen release from pollen (pollen potency) was much higher in Córdoba (3.9 pg Ole e 1/pollen) than in Évora (0.8 pg Ole e 1/pollen, P = 0.004). Indeed, yearly olive pollen counts in Córdoba were 2.4 times higher than in Évora, but Ole e 1 concentrations were 7.6 times higher. When modeling the origin of the pollen, >40% of Ole e 1 exposure in Évora was explained by high-potency pollen originating from the south of Spain. Thus, olive pollen can vary substantially in allergen release, even though they are morphologically identical.

Adhesion to the yeast cell surface as a mechanism for trapping pathogenic bacteria by Saccharomyces probiotics.

Recently, much attention has been given to the use of probiotics as an adjuvant for the prevention or treatment of gastrointestinal pathology. The great advantage of therapy with probiotics is that they have few side effects such as selection of resistant bacteria or disturbance of the intestinal microbiota, which occur when antibiotics are used. Adhesion of pathogenic bacteria onto the surface of probiotics instead of onto intestinal receptors could explain part of the probiotic effect. Thus, this study evaluated the adhesion of pathogenic bacteria onto the cell wall of Saccharomyces boulardii and Saccharomyces cerevisiae strains UFMG 905, W303 and BY4741. To understand the mechanism of adhesion of pathogens to yeast, cell-wall mutants of the parental strain of Saccharomyces cerevisiae BY4741 were used because of the difficulty of mutating polyploid yeast, as is the case for Saccharomyces cerevisiae and Saccharomyces boulardii. The tests of adhesion showed that, among 11 enteropathogenic bacteria tested, only Escherichia coli, Salmonella Typhimurium and Salmonella Typhi adhered to the surface of Saccharomyces boulardii, Saccharomyces cerevisiae UFMG 905 and Saccharomyces cerevisiae BY4741. The presence of mannose, and to some extent bile salts, inhibited this adhesion, which was not dependent on yeast viability. Among 44 cell-wall mutants of Saccharomyces cerevisiae BY4741, five lost the ability to fix the bacteria. Electron microscopy showed that the phenomenon of yeast-bacteria adhesion occurred both in vitro and in vivo (in the digestive tract of dixenic mice). In conclusion, some pathogenic bacteria were captured on the surface of Saccharomyces boulardii, Saccharomyces cerevisiae UFMG 905 and Saccharomyces cerevisiae BY4741, thus preventing their adhesion to specific receptors on the intestinal epithelium and their subsequent invasion of the host.

Isolation of high quality and polysaccharide-free DNA from leaves of Dimorphandra mollis (Leguminosae), a tree from the Brazilian Cerrado.

Dimorphandra mollis (Leguminosae), known as faveiro and fava d'anta, is a tree that is widely distributed throughout the Brazilian Cerrado (a savanna-like biome). This species is economically valuable and has been extensively exploited because its fruits contain the flavonoid rutin, which is used to produce medications for human circulatory diseases. Knowledge about its genetic diversity is needed to guide decisions about the conservation and rational use of this species in order to maintain its diversity. DNA extraction is an essential step for obtaining good results in a molecular analysis. However, DNA isolation from plants is usually compromised by excessive contamination by secondary metabolites. DNA extraction of D. mollis, mainly from mature leaves, results in a highly viscous mass that is difficult to handle and use in techniques that require pure DNA. We tested four protocols for plant DNA extraction that can be used to minimize problems such as contamination by polysaccharides, which is more pronounced in material from mature leaves. The protocol that produced the best DNA quality initially utilizes a sorbitol buffer to remove mucilaginous polysaccharides. The macerated leaf material is washed with this buffer until there is no visible mucilage in the sample. This protocol is adequate for DNA extraction both from young and mature leaves, and could be useful not only for D. mollis but also for other species that have high levels of polysaccharide contamination during the extraction process.

The involvement of calcium carriers and of the vacuole in the glucose-induced calcium signaling and activation of the plasma membrane H(+)-ATPase in Saccharomyces cerevisiae cells.

Previous work from our laboratories demonstrated that the sugar-induced activation of plasma membrane H(+)-ATPase in Saccharomyces cerevisiae is dependent on calcium metabolism with the contribution of calcium influx from external medium. Our results demonstrate that a glucose-induced calcium (GIC) transporter, a new and still unidentified calcium carrier, sensitive to nifedipine and gadolinium and activated by glucose addition, seems to be partially involved in the glucose-induced activation of the plasma membrane H(+)-ATPase. On the other hand, the importance of calcium carriers that can release calcium from internal stores was analyzed in glucose-induced calcium signaling and activation of plasma membrane H(+)-ATPase, in experimental conditions presenting very low external calcium concentrations. Therefore the aim was also to investigate how the vacuole, through the participation of both Ca(2+)-ATPase Pmc1 and the TRP homologue calcium channel Yvc1 (respectively, encoded by the genes PMC1 and YVC1) contributes to control the intracellular calcium availability and the plasma membrane H(+)-ATPase activation in response to glucose. In strains presenting a single deletion in YVC1 gene or a double deletion in YVC1 and PMC1 genes, both glucose-induced calcium signaling and activation of the H(+)-ATPase are nearly abolished. These results suggest that Yvc1 calcium channel is an important component of this signal transduction pathway activated in response to glucose addition. We also found that by a still undefined mechanism Yvc1 activation seems to correlate with the changes in the intracellular level of IP(3). Taken together, these data demonstrate that glucose addition to yeast cells exposed to low external calcium concentrations affects calcium uptake and the activity of the vacuolar calcium channel Yvc1, contributing to the occurrence of calcium signaling connected to plasma membrane H(+)-ATPase activation.

Detection of two morphologically cryptic species from the cursor complex (Akodon spp; Rodentia, Cricetidae) through the use of RAPD markers.

The cursor complex is a group within the Akodon genus of South American rodents, formed by Akodon cursor and A. montensis. Correct distinction between these two species is of great importance since they can harbor different Hantavirus strains. These species are only distinguishable by means of karyotypic or internal anatomic features, requiring dissection; recently, some other genetic methods have become available. We developed RAPD markers capable of distinguishing between A. cursor and A. montensis. Samples included 42 individuals of A. cursor from four localities and 16 individuals of A. montensis from two localities. Fifty-five bands, 41 of which were polymorphic, were analyzed. A principal component analysis showed that this set of markers could successfully distinguish between the two species, mainly based on three RAPD bands. The number of bands in each population was compared within a 95% confidence interval as a measure of intraspecific variability. The A. cursor populations were found to have marked genetic structure across the study area (AMOVA; F(ST )= 0.21), which in part might be because of the relatively limited dispersal capabilities of this species. Species-specific bands, with potential for species identification, were identified.

Evaluation of miltefosine for the treatment of dogs naturally infected with L. infantum (=L. chagasi) in Brazil.

Dogs naturally infected with Leishmania Infantum (=L. chagasi) were treated with miltefosine using different therapeutic regimens. The animals were evaluated for clinical evolution, biochemical parameters, parasite load (by real-time PCR), cytokine levels and humoral response. After treatment and during the following 24 months, there was progressive clinical improvement and complete recovery in 50% (7/14) of the treated animals. There was a decrease in the smear positivity of the bone marrow after treatment, and there was also a gradual and constant decrease in positive cultures at the end of the follow-up period. However, the PCR detection of parasite DNA remained positive. In general, all animals presented a significant increase in parasite load 6 months after treatment. The IFN-γ levels in all the groups tended to increase during follow-up period, regardless of the miltefosine dose administered. The IL-4 and IL-10 levels of the animals tended to decrease during follow-up, except after 300 days when only IL-10 increased. The serum antibodies identified antigens that ranged from 116 kDa to less than 29 kDa in the Western blot assay. Furthermore, 300 days after treatment, qualitative and quantitative differences in the antigen profiles were observed. Antigens of 97 and 46 kDa were the most intensely recognized. Higher levels of antigen-specific Leishmania IgG were detected before and 300 days after treatment in all groups. Taking together, the improvement in the clinical symptoms was not followed by parasitological clearance, suggesting that treatment with miltefosine is not recommended, especially in endemic areas like Brazil, where children are the major victims and dogs are involved in the maintenance of the parasite cycle.

Ocorrência de spalangia endius Walker, 1839 (Hymenoptera, Pteromalidae) em pupas de musca domestica L. e Stomoxys calcitrans L. (Diptera, Muscidae) no sul do Rio Grande do Sul / Occurrence of spalangia endius Walker, 1839 (Hymenoptera, Pteromalidae) in pupae of Musca domestica L. and Stomoxys calcitrans L. in southern Rio Grande do Sul, Brazil

It was reported the occurrence of Spalangia endius Walker, 1839 (Hymenoptera, Pteromalidae) as a parasitoid of pupae of Musca domestica Linnaeus, 1758 (Diptera, Muscidae) and Stomoxys calcitrans Linnaeus, 1758 (Diptera, Muscidae) in the extreme Southern of Brazil. The collection of pupae was performed in January and February, 2008. The pupae of M. domestica and S. calcitrans were collected from bovine feces using the flotation method. The pupae were individualized in glass tubes and maintained in acclimatized chamber at 27±2ºC with relative air humidity > 70 percent until the emergence of the flies or the parasitoids. The referred occurrence consists in the first report to Rio Grande do Sul.

Genomics and proteomics approaches to the study of cancer-stroma interactions.

BACKGROUND: The development and progression of cancer depend on its genetic characteristics as well as on the interactions with its microenvironment. Understanding these interactions may contribute to diagnostic and prognostic evaluations and to the development of new cancer therapies. Aiming to investigate potential mechanisms by which the tumor microenvironment might contribute to a cancer phenotype, we evaluated soluble paracrine factors produced by stromal and neoplastic cells which may influence proliferation and gene and protein expression. METHODS: The study was carried out on the epithelial cancer cell line (Hep-2) and fibroblasts isolated from a primary oral cancer. We combined a conditioned-medium technique with subtraction hybridization approach, quantitative PCR and proteomics, in order to evaluate gene and protein expression influenced by soluble paracrine factors produced by stromal and neoplastic cells. RESULTS: We observed that conditioned medium from fibroblast cultures (FCM) inhibited proliferation and induced apoptosis in Hep-2 cells. In neoplastic cells, 41 genes and 5 proteins exhibited changes in expression levels in response to FCM and, in fibroblasts, 17 genes and 2 proteins showed down-regulation in response to conditioned medium from Hep-2 cells (HCM). Nine genes were selected and the expression results of 6 down-regulated genes (ARID4A, CALR, GNB2L1, RNF10, SQSTM1, USP9X) were validated by real time PCR. CONCLUSIONS: A significant and common denominator in the results was the potential induction of signaling changes associated with immune or inflammatory response in the absence of a specific protein.

Real-time PCR in clinical practice: a powerful tool for evaluating Leishmania chagasi loads in naturally infected dogs.

The performance of the less expensive SYBR-Green-based PCR assay, for quantifying Leishmania chagasi in smears of bone-marrow aspirates from naturally infected, mongrel dogs, was recently compared with that of a similar PCR based on TaqMan chemistry. Aspirates were obtained from 36 infected dogs and examined for parasites by direct examination, culture, and quantitative PCR (qPCR) using specific primers (based on the parasite's kinetoplast DNA), DNA extracted from a smear, and either the SYBR-Green or TaqMan chemistries. Every aspirate smear was found PCR-positive for L. chagasi (whether the assay employed SYBR Green or TaqMan) but only 74% of the aspirates were found positive by culture and only 33% by direct, microscopical examination. There was no evidence of PCR inhibition when the DNA was collected from smears, and the parasite loads estimated using the SYBR-Green PCR were almost identical to those estimated using the TaqMan PCR (r=0.99). As a method for quantifying parasite loads in dogs infected with L. chagasi (and, probably, other mammals infected with other leishmanial parasites), PCR based on SYBR Green may therefore be an appropriate and inexpensive alternative to PCR based on TaqMan, and a reliable clinical tool.