p.Q192R SNP of PON1 seems not to be Associated with Carotid Atherosclerosis Risk Factors in an Asymptomatic and Normolipidemic Brazilian Population Sample

Background:Evidences suggest that paraoxonase 1 (PON1) confers important antioxidant and anti-inflammatory properties when associated with high-density lipoprotein (HDL).Objective:To investigate the relationships between p.Q192R SNP ofPON1, biochemical parameters and carotid atherosclerosis in an asymptomatic, normolipidemic Brazilian population sample.Methods:We studied 584 volunteers (females n = 326, males n = 258; 19-75 years of age). Total genomic DNA was extracted and SNP was detected in the TaqMan® SNP OpenArray® genotyping platform (Applied Biosystems, Foster City, CA). Plasma lipoproteins and apolipoproteins were determined and PON1 activity was measured using paraoxon as a substrate. High-resolution β-mode ultrasonography was used to measure cIMT and the presence of carotid atherosclerotic plaques in a subgroup of individuals (n = 317).Results:The presence of p.192Q was associated with a significant increase in PON1 activity (RR = 12.30 (11.38); RQ = 46.96 (22.35); QQ = 85.35 (24.83) μmol/min; p < 0.0001), HDL-C (RR= 45 (37); RQ = 62 (39); QQ = 69 (29) mg/dL; p < 0.001) and apo A-I (RR = 140.76 ± 36.39; RQ = 147.62 ± 36.92; QQ = 147.49 ± 36.65 mg/dL; p = 0.019). Stepwise regression analysis revealed that heterozygous and p.192Q carriers influenced by 58% PON1 activity towards paraoxon. The univariate linear regression analysis demonstrated that p.Q192R SNP was not associated with mean cIMT; as a result, in the multiple regression analysis, no variables were selected with 5% significance. In logistic regression analysis, the studied parameters were not associated with the presence of carotid plaques.Conclusion:In low-risk individuals, the presence of the p.192Q variant ofPON1 is associated with a beneficial plasma lipid profile but not with carotid atherosclerosis.

A simplified method for sample collection and DNA isolation for polymerase chain reaction detection of Trypanosoma rangeli and Trypanosoma cruzi in triatomine vectors

Due to the overlapping distribution of Trypanosoma rangeli and T. cruzi in Central and South America, sharing several reservoirs and triatomine vectors, we herein describe a simple method to collect triatomine feces and hemolymph in filter paper for further detection and specific characterization of these two trypanosomes. Experimentally infected triatomines feces and hemolymph were collected in filter paper and specific detection of T. rangeli or T. cruzi DNA by polymerase chain reaction was achieved. This simple DNA collection method allows sample collection in the field and further specific trypanosome detection and characterization in the laboratory.

The sample processing time interval as an influential factor in flow cytometry analysis of lymphocyte subsets

The objective of this paper is to propose a protocol to analyze blood samples in yellow fever 17DD vaccinated which developed serious adverse events. We investigated whether or not the time between sample collection and sample processing could interfere in lymphocyte subset percentage, for it is often impossible to analyze blood samples immediately after collection due to transport delay from collection places to the flow cytometry facility. CD4+CD38+ T, CD8+CD38+ T, CD3+ T, CD19+ B lymphocyte subsets were analyzed by flow cytometry in nine healthy volunteers immediately after blood collection and after intervals of 24 and 48 h. The whole blood lysis method and gradient sedimentation by Histopaque were applied to isolate peripheral blood mononuclear cells for flow cytometry analyses. With the lysis method, there was no significant change in lymphocyte subset percentage between the two time intervals (24 and 48 h). In contrast, when blood samples were processed by Histopaque gradient sedimentation, time intervals for sample processing influenced the percentage in T lymphocyte subsets but not in B cells. From the results obtained, we could conclude that the whole blood lysis method is more appropriate than gradient sedimentation by Histopaque for immunophenotyping of blood samples collected after serious adverse events, due to less variation in the lymphocyte subset levels with respect to the time factor.

Paleoparasitological report on Ascaris aDNA from an ancient East Asian sample

In this study, Ascaris DNA was extracted and sequenced from a medieval archaeological sample in Korea. While Ascaris eggs were confirmed to be of human origin by archaeological evidence, it was not possible to pinpoint the exact species due to close genetic relationships among them. Despite this shortcoming, this is the first Ascaris ancient DNA (aDNA) report from a medieval Asian country and thus will expand the scope of Ascaris aDNA research.

Effect of serum sample inactivation on the performance of latex agglutination test for paracoccidioidomycosis serodiagnosis

Paracoccidioidomycosis is diagnosed from the direct observation of the causative agent, but serology can facilitate and decrease the time required for diagnosis. The objective of this study was to determine the influence of serum sample inactivation on the performance of the latex agglutination test (LAT) for detecting antibodies against Paracoccidioides brasiliensis. The sensitivity of LAT from inactivated or non-inactivated samples was 73% and 83%, respectively and the LAT selectivity was 79% and 90%, respectively. The LAT evaluated here was no more specific than the double-immunodiffusion assay. We suggest the investigation of other methods for improving the LAT, such as the use of deglycosylated antigen.

Can Mosquito Magnet® substitute for human-landing catches to sample anopheline populations?

The efficiency of the Mosquito Magnet Liberty PlusTM (MMLP) trap was evaluated in comparison to human-landing catches (HLCs) to sample anopheline populations in Jabillal, state of Bolivar, southern Venezuela. The village comprised 37 houses and a population of 101; malaria in this village is primarily due to Plasmodium vivax and the Annual Parasite Index is 316.8 per 1,000 population. A longitudinal study was conducted between June 2008-January 2009 for three nights per month every two months between 17:30 pm-21:30 pm, a time when biting mosquitoes are most active. Anopheles darlingi and Anopheles nuneztovari were the most common species collected by both methods, whereas Anopheles marajoara was more abundant according to the HLC method. The MMLP trap was more efficient for collecting An. nuneztovari [63%, confidence interval (CI): 2.53] than for collecting An. darlingi (31%, CI: 1.57). There were significant correlations (p < 0.01) between the two methods for An. darlingi [Pearson correlation (R²) = 0.65] and An. nuneztovari (R² = 0.48). These preliminary results are encouraging for further investigations of the use of the MMLP trap for monitoring anopheline populations in remote malaria-endemic areas in the Amazon Basin.

Stool sample storage conditions for the preservation of Giardia intestinalis DNA

Stool is chemically complex and the extraction of DNA from stool samples is extremely difficult. Haemoglobin breakdown products, such as bilirubin, bile acids and mineral ions, that are present in the stool samples, can inhibit DNA amplification and cause molecular assays to produce false-negative results. Therefore, stool storage conditions are highly important for the diagnosis of intestinal parasites and other microorganisms through molecular approaches. In the current study, stool samples that were positive for Giardia intestinalis were collected from five different patients. Each sample was stored using one out of six different storage conditions [room temperature (RT), +4ºC, -20ºC, 70% alcohol, 10% formaldehyde or 2.5% potassium dichromate] for DNA extraction procedures at one, two, three and four weeks. A modified QIAamp Stool Mini Kit procedure was used to isolate the DNA from stored samples. After DNA isolation, polymerase chain reaction (PCR) amplification was performed using primers that target the β-giardin gene. A G. intestinalis-specific 384 bp band was obtained from all of the cyst-containing stool samples that were stored at RT, +4ºC and -20ºC and in 70% alcohol and 2.5% potassium dichromate; however, this band was not produced by samples that had been stored in 10% formaldehyde. Moreover, for the stool samples containing trophozoites, the same G. intestinalis-specific band was only obtained from the samples that were stored in 2.5% potassium dichromate for up to one month. As a result, it appears evident that the most suitable storage condition for stool samples to permit the isolation of G. intestinalis DNA is in 2.5% potassium dichromate; under these conditions, stool samples may be stored for one month.

Association of the solute carrier family 11 member 1 gene polymorphisms with susceptibility to leprosy in a Brazilian sample

Natural resistance-associated macrophage protein 1/solute carrier family 11 member 1 gene (Nramp1/Slc11a1) is a gene that controls the susceptibility of inbred mice to intracellular pathogens. Polymorphisms in the human Slc11a1/Nramp1 gene have been associated with host susceptibility to leprosy. This study has evaluated nine polymorphisms of the Slc11a1/Nramp1 gene [(GT)n, 274C/T, 469+14G/C, 577-18G/A, 823C/T, 1029 C/T, 1465-85G/A, 1703G/A, and 1729+55del4] in 86 leprosy patients (67 and 19 patients had the multibacillary and the paucibacillary clinical forms of the disease, respectively), and 239 healthy controls matched by age, gender, and ethnicity. The frequency of allele 2 of the (GT)n polymorphism was higher in leprosy patients [p = 0.04, odds ratio (OR) = 1.49], whereas the frequency of allele 3 was higher in the control group (p = 0.03; OR = 0.66). Patients carrying the 274T allele (p = 0.04; OR = 1.49) and TT homozygosis (p = 0.02; OR = 2.46), such as the 469+14C allele (p = 0.03; OR = 1.53) of the 274C/T and 469+14G/C polymorphisms, respectively, were more frequent in the leprosy group. The leprosy and control groups had similar frequency of the 577-18G/A, 823C/T, 1029C/T, 1465-85G/A, 1703G/A, and 1729+55del4 polymorphisms. The 274C/T polymorphism in exon 3 and the 469+14G/C polymorphism in intron 4 were associated with susceptibility to leprosy, while the allele 2 and 3 of the (GT)n polymorphism in the promoter region were associated with susceptibility and protection to leprosy, respectively.

Evaluation of sample preparation methods and optimization of nickel determination in vegetable tissues

Nickel, although essential to plants, may be toxic to plants and animals. It is mainly assimilated by food ingestion. However, information about the average levels of elements (including Ni) in edible vegetables from different regions is still scarce in Brazil. The objectives of this study were to: (a) evaluate and optimize a method for preparation of vegetable tissue samples for Ni determination; (b) optimize the analytical procedures for determination by Flame Atomic Absorption Spectrometry (FAAS) and by Electrothermal Atomic Absorption (ETAAS) in vegetable samples and (c) determine the Ni concentration in vegetables consumed in the cities of Lorena and Taubaté in the Vale do Paraíba, State of São Paulo, Brazil. By means of the analytical technique for determination by ETAAS or FAAS, the results were validated by the test of analyte addition and recovery. The most viable method tested for quantification of this element was HClO4-HNO3 wet digestion. All samples but carrot tissue collected in Lorena contained Ni levels above the permitted by the Brazilian Ministry of Health. The most disturbing results, requiring more detailed studies, were the Ni concentrations measured in carrot samples from Taubaté, where levels were five times higher than permitted by Brazilian regulations.

USE OF SCALED SEMIVARIOGRAMS IN THE PLANNING SAMPLE OF SOIL PHYSICAL PROPERTIES IN SOUTHERN AMAZONAS, BRAZIL

There is a great lack of information from soil surveys in the southern part of the State of Amazonas, Brazil. The use of tools such as geostatistics may improve environmental planning, use and management. In this study, we aimed to use scaled semivariograms in sample design of soil physical properties of some environments in Amazonas. We selected five areas located in the south of the state of Amazonas, Brazil, with varied soil uses, such as forest, archaeological dark earth (ADE), pasture, sugarcane cropping, and agroforestry. Regular mesh grids were set up in these areas with 64 sample points spaced at 10 m from each other. At these points, we determined the particle size composition, soil resistance to penetration, moisture, soil bulk density and particle density, macroporosity, microporosity, total porosity, and aggregate stability in water at a depth of 0.00-0.20 m. Descriptive and geostatistical analyses were performed. The sample density requirements were lower in the pasture area but higher in the forest. We concluded that managed-environments had differences in their soil physical properties compared to the natural forest; notably, the soil in the ADE environment is physically improved in relation to the others. The physical properties evaluated showed a structure of spatial dependence with a slight variability of the forest compared to the others. The use of the range parameter of the semivariogram analysis proved to be effective in determining an ideal sample density.

USE OF SCALED SEMIVARIOGRAMS IN THE PLANNING SAMPLE OF SOIL CHEMICAL PROPERTIES IN SOUTHERN AMAZONAS, BRAZIL

The lack of information concerning the variability of soil properties has been a major concern of researchers in the Amazon region. Thus, the aim of this study was to evaluate the spatial variability of soil chemical properties and determine minimal sampling density to characterize the variability of these properties in five environments located in the south of the State of Amazonas, Brazil. The five environments were archaeological dark earth (ADE), forest, pasture land, agroforestry operation, and sugarcane crop. Regular 70 × 70 m mesh grids were set up in these areas, with 64 sample points spaced at 10 m distance. Soil samples were collected at the 0.0-0.1 m depth. The chemical properties of pH in water, OM, P, K, Ca, Mg, H+Al, SB, CEC, and V were determined at these points. Data were analyzed by descriptive and geostatistical analyses. A large part of the data analyzed showed spatial dependence. Chemical properties were best fitted to the spherical model in almost all the environments evaluated, except for the sugarcane field with a better fit to the exponential model. ADE and sugarcane areas had greater heterogeneity of soil chemical properties, showing a greater range and higher sampling density; however, forest and agroforestry areas had less variability of chemical properties.

Can Soil Penetration Resistance and Bulk Density Be Determined in a Single Undisturbed Sample?

Soil quality indicators such as penetration resistance (PR) and bulk density (BD) are traditionally determined in a single undisturbed soil sample. The aim of this study was to assess the effect of PR measurements of undisturbed samples on the determination of BD in the same sample of two soils differing in clay contents. To this end, samples were collected from the 0.00-0.10 and 0.10-0.20 m layers of two soils of clayey and very clayey texture. Volumetric rings were used to collect a total of 120 undisturbed soil samples from each soil layer that were divided into two subsets containing 60 units each. One sample set, designated “perforated samples”, was used to determine PR and BD in the same undisturbed sample; the other, named “intact samples”, was used to determine BD only. Bulk density values for perforated and intact samples were compared by analysis of variance, using a completely randomized experimental design. Means were compared by the t-test at 5 %. The BD values for the clayey soil were similar in perforated and intact samples from the two layers. However, BD of the very clayey soil was lower in the perforated than in the intact samples at both depths. Therefore, PR and BD in clayey soils can be accurately determined in the same undisturbed sample whereas in very clayey soils, different samples are required for this purpose.

Queijo gorgonzola fabricado com leite pasteurizado por ejetor de vapor e HTST: parâmetros físico-químicos e sensoriais

Este trabalho teve como objetivo avaliar a influência de dois sistemas de pasteurização (High Temperature Short Time ¾ HTST¾ e ejetor de vapor) nas características físico-químicas e sensoriais do queijo tipo Gorgonzola. As coletas de amostras de queijo e análises foram realizadas aos 5, 25, 45, 65 e 85 dias de maturação. Durante o período de maturação ocorreu aumento gradual de pH, sal/umidade e índice de acidez nos dois tratamentos. Os queijos fabricados com leite pasteurizado pelo sistema HTST obtiveram valores médios de pH superiores aos dos queijos fabricados com leite pasteurizado pelo sistema ejetor de vapor; os queijos fabricados com leite pasteurizado pelo sistema ejetor de vapor, obtiveram teores de índice de acidez e metilcetonas superiores aos dos queijos fabricados com leite pasteurizado pelo sistema HTST, indicando maior atividade lipolítica nesses queijos. Pela análise sensorial realizada aos 65 dias de maturação, pode se observar que não houve diferença significativa entre os tratamentos em relação à aparência, cor, consistência, textura e sabor. Porém foi observada diferença significativa com relação ao desenvolvimento do mofo e aroma. Os queijos fabricados com leite pasteurizado pelo sistema ejetor de vapor apresentaram maior atividade lipolítica e massa mais macia e fechada.

Sample size for measurement of root traits on common bean by image analysis

Evaluation of root traits may be facilitated if they are assessed on samples of the root system. The objective of this work was to determine the sample size of the root system in order to estimate root traits of common bean (Phaseolus vulgaris L.) cultivars by digital image analysis. One plant was grown per pot and harvested at pod setting, with 64 and 16 pots corresponding to two and four cultivars in the first and second experiments, respectively. Root samples were scanned up to the completeness of the root system and the root area and length were estimated. Scanning a root sample demanded 21 minutes, and scanning the entire root system demanded 4 hours and 53 minutes. In the first experiment, root area and length estimated with two samples showed, respectively, a correlation of 0.977 and 0.860, with these traits measured in the entire root. In the second experiment, the correlation was 0.889 and 0.915. The increase in the correlation with more than two samples was negligible. The two samples corresponded to 13.4% and 16.9% of total root mass (excluding taproot and nodules) in the first and second experiments. Taproot stands for a high proportion of root mass and must be deducted on root trait estimations. Samples with nearly 15% of total root mass produce reliable root trait estimates.

Sample size for full-sib family evaluation in sugarcane

The objective of this study was to determine the minimum number of plants per plot that must be sampled in experiments with sugarcane (Saccharum officinarum) full-sib families in order to provide an effective estimation of genetic and phenotypic parameters of yield-related traits. The data were collected in a randomized complete block design with 18 sugarcane full-sib families and 6 replicates, with 20 plants per plot. The sample size was determined using resampling techniques with replacement, followed by an estimation of genetic and phenotypic parameters. Sample-size estimates varied according to the evaluated parameter and trait. The resampling method permits an efficient comparison of the sample-size effects on the estimation of genetic and phenotypic parameters. A sample of 16 plants per plot, or 96 individuals per family, was sufficient to obtain good estimates for all traits considered of all the characters evaluated. However, for Brix, if sample separation by trait were possible, ten plants per plot would give an efficient estimate for most of the characters evaluated.