Sequence and transcript expression of the super-kdr locus of the horn fly, Haematobia irritans.

In horn flies, Haematobia irritans irritans (Diptera: Muscidae) (Linnaeus, 1758), target site resistance to pyrethroids can be diagnosed by an allele-specific PCR that genotypes individual flies at both the super-kdr (skdr) and the knock down resistance (kdr) associated loci. When this technique uses genomic DNA as template, modifications, such as alternative RNA splicing and RNA editing are not specifically detected. Alternative splicing at the skdr locus has been reported in Dipterans; thus, the genomic DNA-based allele-specific PCR may not accurately reflect the frequency of the skdr mutation in horn fly field populations. To investigate if alternative splicing occurs at the skdr locus of horn flies, genomic DNA and cDNA sequences isolated from two wild populations and two laboratory-reared colonies with varying degrees of pyrethroid resistance were compared. There was no indication of alternative splicing at the super-kdr locus neither in the wild populations nor in the laboratory-reared colonies.

Seroepidemiology of Bovine Herpes Virus-1 Infection in Water Buffaloes from the state of Veracruz, Mexico.

Water buffaloes (Bubalus bubalis) were introduced to Mexico at the end of the last century. In Mexico, buffaloes are commonly pastured together with cattle; however few studies have been done on buffalo herd health in Mexico. We hypothesized that a better knowledge of the epidemiology of infections shared between cattle and buffaloes may improve herd profitability and promote buffalo production in areas unsuitable to cattle farming. This study aimed to determine the prevalence of antibodies against bovine herpes virus - 1 (BoHV1) in water buffaloes raised on six farms from the state of Veracruz, Mexico. Of 368 buffaloes sampled, 217 (59%) were seropositive for BoHV-1. Age was identified as a risk factor for BoHV-1 infection with buffaloes older than 5 years being the most likely to be infected. Animals more than 7 years old had the highest prevalence (86.0%). Females and males had similar seroprevalence rates. Females with history of abortion had higher prevalence of antiBoHV-1 antibodies than those with no record of abortion. Buffaloes and cattle were raised together in only one of the six farms under study. Interaction with cattle was not a risk factor for BoHV-1 seropositivity. This study showed that BoHV-1 is prevalent among buffalo herds in the state of Veracruz, Mexico. Buffaloes appear to play an important role in the epidemiology of BoHV-1 infection in parts of Mexico when there is no apparent risk of interaction with cattle. Animal health programs established to mitigate the burden caused by BoHV-1 must take into consideration buffaloes when this bovid species is part of the agroecosystem shared with cattle.

Effect of ultrasonic and sonic activation of root canal sealers on the push-out bond strength and interfacial adaptation to root canal dentine.

AIM: To evaluate the effect of ultrasonic and sonic activation of two root canal sealers on interfacial adaptation and push-out bond strength (BS) to root canal dentine. METHODOLOGY: The root canals of seventy-eight roots of maxillary canines were prepared with NiTi rotary instruments under irrigation with 2.5% NaOCl and 17% EDTA. Canals were filled by lateral condensation of gutta-percha and AH Plus (n = 39) or MTA Fillapex (n = 39). Three subgroups (n = 13) were formed according to sealer activation: no activation (NA), sonic activation (SA, 20 s) and ultrasonic activation (US, 20 s). In three specimens of each subgroup, 0.1% rhodamine B was added to the sealer. Three 1-mm-thick slices were obtained from each root third. The first and second were used for push-out BS test and the third for a qualitative analysis of interfacial adaptation (gaps) and voids by confocal laser scanning microscopy (CLSM). Intratubular penetration of rhodamine B-labelled sealer was also assessed by CLSM. Data were analysed by two-way anova and Tukey's test (α = 0.05). RESULTS: AH Plus (2.58 ± 1.21) had higher push-out BS than MTA Fillapex (1.45 ± 0.71) (P = 0.000). UA (2.64 ± 1.44) had higher BS (P = 0.000) than NA (1.58 ± 0.73) and SA (1.83 ± 0.85), which did not differ significantly from each other (P = 0.3303). The coronal third had higher BS (2.48 ± 1.49) than the middle (2.08 ± 0.94) (P = 0.0003) and apical (1.48 ± 0.58) (P = 0.00) thirds. For both sealers, UA was associated with a more homogeneous filling material with better interfacial adaptation and greater quantity, density and length of tags in dentinal tubules. When UA was used, the tags had similar quantity, density and length to those formed without activation, but interfacial gaps and voids were observed. CONCLUSION: Ultrasonic activation was associated with higher BS values, deeper intratubular penetration and greater interfacial adaptation to root dentine than sonic activation and no activation techniques, AH Plus had higher BS values than MTA Fillapex irrespective of the type of activation.

SCN9A channelopathy associated autosomal recessive Congenital Indifference to Pain. A case report.

BACKGROUND: Congenital Indifference to Pain (CIP) is a rare condition that inhibits the ability of patients to perceive physical pain but otherwise keeps normal sensory modalities. The condition has been mapped to an autosomal recessive trait to chromosome 2q 24.3 with mutations on the SCN9A gene. CASE REPORT: A 2 year old Caucasian female presented with CIP. Bite injuries, tongue wounds and unaccounted dental trauma episodes were frequently reported. Preventive instructions and possible treatment modalities were discussed with the parents. CONCLUSION: The cornerstone of treating CIP patients is an extensive preventive approach alongside regular oral examination at home by parents as well as routine recall appointments with dentists.

Drought sensitivity of Amazonian carbon balance revealed by atmospheric measurements.

Feedbacks between land carbon pools and climate provide one of the largest sources of uncertainty in our predictions of global climate. Estimates of the sensitivity of the terrestrial carbon budget to climate anomalies in the tropics and the identification of the mechanisms responsible for feedback effects remain uncertain. The Amazon basin stores a vast amount of carbon, and has experienced increasingly higher temperatures and more frequent floods and droughts over the past two decades. Here we report seasonal and annual carbon balances across the Amazon basin, based on carbon dioxide and carbon monoxide measurements for the anomalously dry and wet years 2010 and 2011, respectively. We find that the Amazon basin lost 0.48 ± 0.18 petagrams of carbon per year (Pg C yr(-1)) during the dry year but was carbon neutral (0.06 ± 0.1 Pg C yr(-1)) during the wet year. Taking into account carbon losses from fire by using carbon monoxide measurements, we derived the basin net biome exchange (that is, the carbon flux between the non-burned forest and the atmosphere) revealing that during the dry year, vegetation was carbon neutral. During the wet year, vegetation was a net carbon sink of 0.25 ± 0.14 Pg C yr(-1), which is roughly consistent with the mean long-term intact-forest biomass sink of 0.39 ± 0.10 Pg C yr(-1) previously estimated from forest censuses. Observations from Amazonian forest plots suggest the suppression of photosynthesis during drought as the primary cause for the 2010 sink neutralization. Overall, our results suggest that moisture has an important role in determining the Amazonian carbon balance. If the recent trend of increasing precipitation extremes persists, the Amazon may become an increasing carbon source as a result of both emissions from fires and the suppression of net biome exchange by drought.

Improving the affinity of fibroblasts for bacterial cellulose using carbohydrate-binding modules fused to RGD.

The attachment of cells to biomedical materials can be improved by using adhesion sequences, such as Arg-Gly-Asp (RGD), found in several extracellular matrix proteins. In this work, bifunctional recombinant proteins, with a Cellulose-Binding Module (CBM), from the cellulosome of Clostridium thermocellum and cell binding sequences-RGD, GRGDY-were cloned and expressed in E.coli. These RGD-containing cellulose binding proteins were purified and used to coat bacterial cellulose fibres. Its effect on the cell adhesion/biocompatibility properties was tested using a mouse embryo fibroblasts culture. Bacterial cellulose (BC) secreted by Gluconacetobacter xylinus (=Acetobacter xylinum) is a material with unique properties and promising biomedical applications. CBMs adsorbs specifically and tightly on cellulose. Thus, they are a useful tool to address the fused RGD sequence (or other bioactive peptides) to the cellulose surface, in a specific and simple way. Indeed, fibroblasts exhibit improved ability to interact with bacterial cellulose sheets coated with RGD-CBM proteins, as compared with cellulose treated with the CBM, that is, without the adhesion peptide. The effect of the several fusion proteins produced was analyzed.

Susceptibilidade do besouro rola-bosta africano a reguladores de crescimento de insetos / Susceptibility of African dung beetle to insect growth regulators

Verificou-se a ação dos reguladores de crescimento de insetos (IGR), diflubenzuron e methoprene, sobre o besouro rola-bosta africano, Digitonthophagus gazella (Fabricius), um inimigo natural da mosca-dos-chifres, Haematobia irritans irritans (Linnaeus). Casais de besouros foram colocados em baldes contendo terra úmida e alimentados com fezes bovinas contendo 1, 0,5 ou 0,2ppm de diflubenzuron e 0,2ppm de methoprene. Os insetos e sua prole foram recuperados com o auxílio de armadilhas pitfall. Diflubenzuron e methoprene não afetaram a sobrevivência dos adultos inicialmente expostos, mas interferiram na produção de descendentes. Diflubenzuron, nas concentrações de 1 e 0,5ppm, também afetou a duração do ciclo de vida dos besouros. Nenhum dos IGRs alterou a razão sexual dos descendentes obtidos. As concentrações testadas de diflubenzuron mostraram-se moderadamente nocivas ao besouro enquanto methoprene a 0,2ppm mostrou ser pouco nocivo, segundo os critérios da International Organization for Biological Control.

The effects of insect growth regulators (IGR), diflubenzuron and methoprene, on African dung beetle, Digitonthophagus gazella (Fabricius), a natural enemy of the horn fly, Haematobia irritans irritans (Linnaeus), was studied. Beetles were placed in buckets partially filled with humid soil and were fed bovine feces containing 1, 0.5, or 0.2ppm diflubenzuron and 0.2ppm methoprene. Insects and their progenies were recovered by pitfall traps. Diflubenzuron and methoprene did not affect the survival of the adults but reduced their progenies. Diflubenzuron 1 and 0.5ppm also affected the life cicle of the beetles. None of the IGR modified the gender ratio of the progenies. According to the IOBC criteria, diflubenzuron tested concentrations showed to be moderately harmful to the beetles, whereas methoprene 0.2ppm was slightly harmful.

Kinetics of lactose fermentation using a recombinant Saccharomyces cerevisiae strain.

This work presents a multi-route, non-structural kinetic model for interpretation of ethanol fermentation of lactose using a recombinant flocculent Saccharomyces cerevisiae strain expressing both the LAC4 (coding for beta-galactosidase) and LAC12 (coding for lactose permease) genes of Kluyveromyces lactis. In this model, the values of different metabolic pathways are calculated applying a modified Monod equation rate in which the growth rate is proportional to the concentration of a key enzyme controlling the single metabolic pathway. In this study, three main metabolic routes for S. cerevisiae are considered: oxidation of lactose, reduction of lactose (producing ethanol), and oxidation of ethanol. The main bioprocess variables determined experimentally were lactose, ethanol, biomass, and dissolved oxygen concentrations. Parameters of the proposed kinetic model were established by fitting the experimental data obtained in a small lab-scale fermentor with the initial lactose concentrations ranging from 5 g/dm3 to 50 g/dm3. A very good agreement between experimental data and simulated profiles of the main variables (lactose, ethanol, biomass, and dissolved oxygen concentrations) was achieved.

Differential diagnosis and treatment of hyponatremia following pituitary surgery.

Hyponatremia is a frequent occurrence after pituitary surgery, having been described in 9% to 35% of the patients. It is produced by the syndrome of inappropriate secretion of antidiuretic hormone (SIADH) or, more frequently, by the cerebral salt-wasting syndrome (CSWS). The clinical presentation of both syndromes is identical and the differential diagnosis can be difficult. The determination of the volemic state is essential for the diagnosis, since the patients with the SIADH are euvolemic or hypervolemic, while those with CSWS are hypovolemic. Several methods can be used to detect the volemic state in these patients and, among them, the furosemide test can best discriminate between SIADH and CSWS. A furosemide infusion (20 mg) normalizes sodium serum levels in SIADH patients, but not in CSWS patients who remain hyponatremic. The differentiation between the 2 syndromes is clinically relevant since their treatment is antithetical. SIADH patients need liquid restriction of liquids and/or furosemide to reduce the volume of extracellular water, while CSWS patients need volume replacement with sodium supplementation (or fludrocortisone can be a good alternative). The diagnosis and treatment of these syndromes are discussed on the basis of the literature reports.

Construction of a flocculent Saccharomyces cerevisiae strain secreting high levels of Aspergillus niger beta-galactosidase.

A flocculent Saccharomyces cerevisiae strain secreting Aspergillus niger beta-galactosidase activity was constructed by transforming S. cerevisiae NCYC869-A3 strain with plasmid pVK1.1 harboring the A. niger beta-galactosidase gene, lacA, under the control of the ADH1 promoter and terminator. Compared to other recombinant S. cerevisiae strains, this recombinant yeast has higher levels of extracellular beta-galactosidase activity. In shake-flask cultures, the beta-galactosidase activity detected in the supernatant was 20 times higher than that obtained with previously constructed strains (Domingues et al. 2000a). In bioreactor culture, with cheese-whey permeate as substrate, a yield of 878.0 nkat/gsubstrate was obtained. The recombinant strain is an attractive alternative to other fungal beta-galactosidase production systems as the enzyme is produced in a rather pure form. Moreover, the use of flocculating yeast cells allows for enzyme production with high productivity in continuous fermentation systems with facilitated downstream processing.

Alcohol production from cheese whey permeate using genetically modified flocculent yeast cells.

Alcoholic fermentation of cheese whey permeate was investigated using a recombinant flocculating Saccharomyces cerevisiae, expressing the LAC4 (coding for beta-galactosidase) and LAC12 (coding for lactose permease) genes of Kluyveromyces marxianus enabling for lactose metabolization. Data on yeast fermentation and growth on cheese whey permeate from a Portuguese dairy industry is presented. For cheese whey permeate having a lactose concentration of 50 gL(-1), total lactose consumption was observed with a conversion yield of ethanol close to the expected theoretical value. Using a continuously operating 5.5-L bioreactor, ethanol productivity near 10 g L(-1) h(-1) (corresponding to 0.45 h(-1) dilution rate) was obtained, which raises new perspectives for the economic feasibility of whey alcoholic fermentation. The use of 2-times concentrated cheese whey permeate, corresponding to 100 gL(-1) of lactose concentration, was also considered allowing for obtaining a fermentation product with 5% (w/v) alcohol.

Construction of a flocculent brewer's yeast strain secreting Aspergillus niger beta-galactosidase.

One way of improving heterologous protein production is to use high cell density systems, one of the most attractive being the flocculating yeast production system. Also, lactose is available in large amounts as a waste product from cheese production processes. The construction of flocculent and non-flocculent brewer's yeast strains secreting beta-galactosidase and growing on lactose is presented. A plasmid was constructed coding for an extracellular beta-galactosidase of Aspergillus niger and having, as selective marker, the yeast CUP1 gene conferring resistance to copper. This selective marker allows for the transformation of wild-type yeasts. This work represents an important step towards the study of heterologous protein secretion by flocculent cells.

Contamination of a high-cell-density continuous bioreactor.

Continuous fermentations were carried out with a recombinant flocculent Saccharomyces cerevisiae strain in an airlift bioreactor. Once operating under steady state at a dilution rate of 0.45 h(-1), the bioreactor was contaminated with Escherichia coli cells. The faster growing E. coli strain was washed out of the bioreactor and the recombinant, slower growing flocculating S. cerevisiae strain remained as the only species detected in the bioreactor. Flocculation, besides allowing for the realization of high-cell-density systems with corresponding unusual high productivity, may be used as a selective property for controlling some contamination problems associated with prolonged continuous operation.

Tipo e freqüência de alteraçöes dentárias e periodontais em cäes na regiäo de Jaboticabal, SP / Type and frequency of dental diseases and disorders in dogs in the region of Jaboticabal, SP

Realizou-se levantamento das alteraçöes periodontais, dentárias e da oclusäo dental espontâneas em 215 cäes necropsiados ou que morreram por razöes diversas, no Hospital Veterinário da Faculdade de Ciências Agrárias e Veterinárias da UNESP, Jaboticabal. As alteraçöes periodontais, identificadas por exame clínico, foram tabuladas considerando raça, dieta, faixa etária e sexo. Os índices gengival e de placa foram aplicados a todos os animais e análise radiográfica foi realizada quando necessária. O levantamento epidemiológico revelou elevada freqüência de alteraçöes relacionadas à presença de placa bacteriana e de alteraçöes gengivais. Observou-se também agravamento das alteraçöes com o avançar da idade, evidenciado radiograficamente. Os animais alimentados somente com raçäo apresentaram menor freqüência de alteraçöes relacionadas aos índices gengival e de placa, em relaçäo àqueles alimentados com dieta caseira. Näo se observou diferença entre sexos quanto às alteraçöes dentárias

Microbiota saprófita associada à doença periodontal em cäes / Oral flora associated with periodontal disease in dogs

Realizou-se um estudo sobre a microbiota saprófita associada à doença periodontal espontânea em cäes com o objetivo de identificar as bactérias anaeróbias predominantes nas lesöes. Com auxílio de cureta odontológica, amostras colhidas diretamente do espaço subgengival foram semeadas em meio CDC (Central for Disease Control) para anaeróbios e incubadas, em anaerobiose, a 37§C, por sete dias. A caracterizaçäo das colônias foi realizada por meio da morfologia e do teste bioquímico (Sistema API 20A). Identificaram-se os seguintes gêneros: Prevotella spp., Bacteroides spp., Propionibacterium spp., Gemella spp., Actinomyces spp., Eubacterium spp. e Porphyromonas spp

Continuous ethanol fermentation of lactose by a recombinant flocculating Saccharomyces cerevisiae strain.

Alcohol fermentation of lactose was investigated using a recombinant flocculating Saccharomyces cerevisiae, expressing the LAC4 (coding for beta-galactosidase) and LAC12 (coding for lactose permease) genes of Kluyveromyces marxianus. Data on yeast fermentation and growth on a medium containing lactose as the sole carbon source are presented. In the range of studied lactose concentrations, total lactose consumption was observed with a conversion yield of ethanol close to the expected theoretical value. For the continuously operating bioreactor, an ethanol productivity of 11 g L(-1) h(-1) (corresponding to a feed lactose concentration of 50 g L(-1) and a dilution rate of 0.55 h(-1)) was obtained, which is 7 times larger than the continuous conventional systems. The system stability was confirmed by keeping it in operation for 6 months.

Construction of a flocculent Saccharomyces cerevisiae fermenting lactose.

A flocculent Saccharomyces cerevisiae strain with the ability to express both the LAC4 (coding for beta-galactosidase) and LAC12 (coding for lactose permease) genes of Kluyveromyces marxianus was constructed. This recombinant strain is not only able to grow on lactose, but it can also ferment this substrate. To our knowledge this is the first time that a recombinant S. cervisiae has been found to ferment lactose in a way comparable to that of the existing lactose-fermenting yeast strains. Moreover, the flocculating capacity of the strain used in this work gives the process several advantages. On the one hand, it allows for operation in a continuous mode at high cell concentration, thus increasing the system's overall productivity; on the other hand, the biomass concentration in the effluent is reduced, thus decreasing product separation/purification costs.

Cytokine production in acute versus chronic human Schistosomiasis mansoni: the cross-regulatory role of interferon-gamma and interleukin-10 in the responses of peripheral blood mononuclear cells and splenocytes to parasite antigens.

The contribution of interleukin (IL)-10 and interferon (IFN)-gamma to the regulation of type 1 and type 2 cytokine responses was investigated in Brazilians with different clinical forms of schistosomiasis mansoni. Cells from members of a family with acute intestinal schistosomiasis responded to schistosomal soluble egg antigen (SEA) or soluble adult worm antigen preparation (SWAP) with greater amounts of IFN-gamma than did cells from several patients with chronic intestinal schistosomiasis; IL-10 levels were similar. Neutralization of IL-10 had no effect on the SEA-specific IFN-gamma response in patients with acute infection, whereas SWAP-induced IFN-gamma was increased in both groups. Anti-IL-10 also up-regulated SEA-specific IFN-gamma protein and mRNA responses in most splenocyte cultures from hepatosplenic schistosomiasis patients but had no effect on antigen-specific IL-4 or IL-5 production. Neutralization of IFN-gamma resulted in a comparable increase in SWAP-specific IL-10 and IL-5, while IL-4 was not affected. These studies demonstrate that early disease in schistosomiasis is associated with a significant IFN-gamma response and that IL-10 contributes to the suppression of that response during both early and chronic infection.

Transformation of a flocculating Saccharomyces cerevisiae using lithium acetate and pYAC4.

A flocculating yeast Saccharomyces cerevisiae ura3 was transformed by the method based on treatment of intact cells with lithium acetate plus single-stranded carrier DNA using the shuttle vector pYAC4. The transformation efficiency was above 10(3) transformants per microgram of plasmid DNA which is similar to other described yeast transformation systems. Under selective pressure, the transformed cells were stable and maintained the flocculation ability. Thus, this simple transformation system can be used for gene expression studies in flocculating yeasts, overcoming disadvantages of conventional methods such as the spheroplast one.