FALSE-NEGATIVE DENGUE CASES IN RORAIMA, BRAZIL: AN APPROACH REGARDING THE HIGH NUMBER OF NEGATIVE RESULTS BY NS1 AG KITS

Serum samples from 150 NS1-negative (Platelia ELISA) patients presumptively diagnosed with dengue were analyzed by the TaqMan probed real-time reverse transcription PCR (TaqMan qRT-PCR) method. The qRT-PCR positive samples were tested for serotype by semi-nested RT-PCR and a qualitative immunochromatographic assay for IgG and IgM. Molecular detection methods showed 33 (22%) positive samples out of 150 NS1-antigen negative samples. Of these, 72% were collected up to day 2 after the onset of symptoms, when diagnostic sensitivity of NS1-antigen test assays is significantly enhanced. Most of the cases were not characterized as secondary infection. Twenty-eight samples were successfully serotyped, 75% of which for DENV-4, 14% for DENV-2, 7% for DENV-3 and 4% for DENV-1. These findings reaffirm the hyperendemic situation of the state of Roraima and suggest a lower sensitivity of the NS1 test, mainly when DENV-4 is the predominant serotype. Health care providers should therefore be aware of samples tested negative by NS1 antigen assays, especially when clinical symptoms and other laboratory data results show evidence of dengue infection.

Performance of a reciprocal shaker in mechanical dispersion of soil samples for particle-size analysis

The dispersion of the samples in soil particle-size analysis is a fundamental step, which is commonly achieved with a combination of chemical agents and mechanical agitation. The purpose of this study was to evaluate the efficiency of a low-speed reciprocal shaker for the mechanical dispersion of soil samples of different textural classes. The particle size of 61 soil samples was analyzed in four replications, using the pipette method to determine the clay fraction and sieving to determine coarse, fine and total sand fractions. The silt content was obtained by difference. To evaluate the performance, the results of the reciprocal shaker (RSh) were compared with data of the same soil samples available in reports of the Proficiency testing for Soil Analysis Laboratories of the Agronomic Institute of Campinas (Prolab/IAC). The accuracy was analyzed based on the maximum and minimum values defining the confidence intervals for the particle-size fractions of each soil sample. Graphical indicators were also used for data comparison, based on dispersion and linear adjustment. The descriptive statistics indicated predominantly low variability in more than 90 % of the results for sand, medium-textured and clay samples, and for 68 % of the results for heavy clay samples, indicating satisfactory repeatability of measurements with the RSh. Medium variability was frequently associated with silt, followed by the fine sand fraction. The sensitivity analyses indicated an accuracy of 100 % for the three main separates (total sand, silt and clay), in all 52 samples of the textural classes heavy clay, clay and medium. For the nine sand soil samples, the average accuracy was 85.2 %; highest deviations were observed for the silt fraction. In relation to the linear adjustments, the correlation coefficients of 0.93 (silt) or > 0.93 (total sand and clay), as well as the differences between the angular coefficients and the unit < 0.16, indicated a high correlation between the reference data (Prolab/IAC) and results obtained with the RSh. In conclusion, the mechanical dispersion by the reciprocal shaker of soil samples of different textural classes was satisfactory. The results allowed recommending the use of the equipment at low agitation for particle size- analysis. The advantages of this Brazilian apparatus are its low cost, the possibility to simultaneously analyze a great number of samples using ordinary, easily replaceable glass or plastic bottles.

Number of buried seeds and seedlings emergence in cerradão, cerrado and gallery forest soils at Pedregulho, Itirapina (SP), Brazil

This study was carried out at the site named Pedregulho in the "Estação Experimental de Itirapina", Itirapina, state of São Paulo, Brazil. Soil samples were collected in the rainy and in the dry season at three depths in the gallery forest, cerrado and cerradão. The total number of seeds was counted after being separated from the soil by wet sieving and the number of viable seeds was determined by emergence of seedlings in soil samples in a greenhouse. The number of seeds and of emerged seedlings decreases as soil samples become deeper. The number of seeds was much larger in the cerrado in the dry season than in the rainy season, but it was lower in the dry than in the rainy season in the gallery forest. The number of seeds in the cerradão was similar in both seasons. There was a heavy discrepancy between the total number of seeds and that of emerged seedlings.

Distribution of aflatoxin contamination in maize samples

The distribution of the aflatoxin contamination was studied among four maize fractions, separated according to Brazilian grading rules for maize. The fraction that contained fermented, moldy, heated and sprouted grains normally had the highest levels of aflatoxin. However, the fraction contribution to the whole sample contamination level took into account the contamination fraction level and its weight to the whole sample. Considering this, the fraction that contained insect damaged, hollow, up to ¼ fermented and grains damaged by other causes was normally the fraction responsible for the total contamination level in the samples. Nevertheless, the fraction contributions were variable from sample to sample. Therefore, in conclusion, it was not possible to establish a standard behavior for grain fraction-type contribution for different maize lots. The Brazilian grading by qualitative types applied to samples did not show statistic correlation with aflatoxin contamination levels (P<0.05). Two type-1 samples (the best quality type) presented contamination of 380 and 146ng/g. The number of samples with contamination levels above those allowed by Brazilian law (20ng/g) was the same for qualitative types 2, 3, and BS (Below Standard).

Can varying the number of teams in a shift schedule constitute a preventive strategy?

OBJECTIVE: The study examines the implications for shiftworkers of applying different numbers of teams in the organization of shiftwork. METHODS: The participating operators came from five different companies applying continuous shift rotation systems. The companies shared the same product organization and a common corporate culture belonging to the same multinational company. Each company had a shift system consisting of four, five or six teams, with the proportion of shifts outside day work decreasing as the number of teams increased. Questionnaire and documentary data were used as data sources. RESULTS: Operators in systems with additional teams had more daywork but also more irregular working hours due to both overtime and schedule changes. Operators using six teams used fewer social compensation strategies. Operators in four teams were most satisfied with their work hours. Satisfaction with the time available for various social activities outside work varied inconsistently between the groups. CONCLUSIONS: In rotating systems the application of more teams reduces the number of shifts outside day work. This apparent improvement for shiftworkers was counteracted by a concomitant irregularity produced by greater organizational requirements for flexibility. The balance of this interaction was found to have a critical impact on employees.

Entamoeba histolytica: fecal antigen capture immunoassay for the diagnosis of enteric amebiasis by a monoclonal antibody

Amebiasis continues to be of epidemiological importance in underdeveloped countries. Clinical diagnosis and epidemiological setting in a region are based on the fecal microscopic identification of cysts or trophozoites. This procedure requires well trained personnel, is laborious, of low sensitivity and frequently yields false-positives results. The present study was designed to develop an immuno-enzymatic fecal 96 kDa antigen capture test (COPROELISA-Eh) more sensitive and specific than microscopic diagnosis of amebiasis. Triplicates of 177 stool samples processed by the formol-ether concentration method, were defined as positive or negative by three experienced microscopic observers. Another aliquot was submitted to the antigen capture test by a monoclonal antibody against a specific membrane antigen of pathogenic strains of Entamoeba histolytica. Optical densities were interpreted as positive when they exceeded the mean value of negative samples plus two standard deviations. COPROELISA-Eh showed a 94.4% sensitivity, 98.3% specificity, 96.2% positive predictive value and 97.6% negative predictive value for the detection of E. histolytica in feces. COPROELISA-Eh is more sensitive and specific than microscopic examination, does not require specially trained personnel and allows the simultaneous processing of a large number of samples.

The use of oligonucleotide probes for meningococcal serotype characterization

In the present study we examine the potential use of oligonucleotide probes to characterize Neisseria meningitidis serotypes without the use of monoclonal antibodies (MAbs). Antigenic diversity on PorB protein forms the bases of serotyping method. However, the current panel of MAbs underestimated, by at least 50% the PorB variability, presumably because reagents for several PorB variable regions (VRs) are lacking, or because a number of VR variants are not recognized by serotype-defining MAbs12. We analyzed the use of oligonucleotide probes to characterize serotype 10 and serotype 19 of N. meningitidis. The porB gene sequence for the prototype strain of serotype 10 was determined, aligned with 7 other porB sequences from different serotypes, and analysis of individual VRs were performed. The results of DNA probes 21U (VR1-A) and 615U (VR3-B) used against 72 N. meningitidis strains confirm that VR1 type A and VR3 type B encode epitopes for serotype-defined MAbs 19 and 10, respectively. The use of probes for characterizing serotypes possible can type 100% of the PorB VR diversity. It is a simple and rapid method specially useful for analysis of large number of samples.

Comparative analysis of two-dimensional electrophoresis maps (2-DE) of Helicobacter pylori from Brazilian patients with chronic gastritis and duodenal ulcer: a preliminary report

Helicobacter pylori is a bacterium recognized as the major cause of peptic ulcer and chronic gastritis. Recently, a proteome-based approach was developed to investigate pathogenic factors related to H. pylori. In this preliminary study, H. pylori strains were isolated from gastric biopsies of patients with chronic gastritis and duodenal ulcers. A partial proteomic analysis of H. pylori strains was performed by bacterial lyses and proteins were separated by two-dimensional gel electrophoresis (2-DE). A comparative analysis was performed to verify a differential protein expression between these two 2-DE maps. These data should be useful to clarify the role of different proteins related to bacterial pathogenesis. This study will be completed using a larger number of samples and protein identification of H. pylori by MALDI-TOF mass spectrometry.

Hepatitis B virus genotyping among chronic hepatitis B patients with resistance to treatment with lamivudine in the City of Ribeirão Preto, State of São Paulo

INTRODUCTION: Lamivudine is a nucleoside analogue that is used clinically for treating chronic hepatitis B infection. However, the main problem with prolonged use of lamivudine is the development of viral resistance to the treatment. Mutations in the YMDD motif of the hepatitis B virus DNA polymerase gene have been associated with resistance to drug therapy. So far, there have not been many studies in Brazil reporting on genotype-dependent development of resistance to lamivudine. Thus, the aim of the present study was to determine the possible correlation between a certain genotype and increased development of resistance to lamivudine among chronic hepatitis B patients. METHODS: HBV DNA in samples from 50 patients under lamivudine treatment was amplified by means of conventional PCR. Samples were collected at Hospital das Clínicas, FMRP-USP. The products were then sequenced and phylogenetic analysis was performed. RESULTS: Phylogenetic analysis revealed that 29 (58%) patients were infected with genotype D, 20 (40%) with genotype A and one (2%) with genotype F. Mutations in the YMDD motif occurred in 20% of the patients with genotype A and 27.6% of the patients with genotype D. CONCLUSIONS: Despite the small number of samples, our results indicated that mutations in the YMDD motif were 1.38 times more frequent in genotype D than in genotype A.

A SARIMA forecasting model to predict the number of cases of dengue in Campinas, State of São Paulo, Brazil

INTRODUCTION: Forecasting dengue cases in a population by using time-series models can provide useful information that can be used to facilitate the planning of public health interventions. The objective of this article was to develop a forecasting model for dengue incidence in Campinas, southeast Brazil, considering the Box-Jenkins modeling approach. METHODS: The forecasting model for dengue incidence was performed with R software using the seasonal autoregressive integrated moving average (SARIMA) model. We fitted a model based on the reported monthly incidence of dengue from 1998 to 2008, and we validated the model using the data collected between January and December of 2009. RESULTS: SARIMA (2,1,2) (1,1,1)12 was the model with the best fit for data. This model indicated that the number of dengue cases in a given month can be estimated by the number of dengue cases occurring one, two and twelve months prior. The predicted values for 2009 are relatively close to the observed values. CONCLUSIONS: The results of this article indicate that SARIMA models are useful tools for monitoring dengue incidence. We also observe that the SARIMA model is capable of representing with relative precision the number of cases in a next year.

Evaluation of two sweeping methods for estimating the number of immature Aedes aegypti (Diptera: Culicidae) in large containers

Introduction Here, we evaluated sweeping methods used to estimate the number of immature Aedes aegypti in large containers. Methods III/IV instars and pupae at a 9:1 ratio were placed in three types of containers with, each one with three different water levels. Two sweeping methods were tested: water-surface sweeping and five-sweep netting. The data were analyzed using linear regression. Results The five-sweep netting technique was more suitable for drums and water-tanks, while the water-surface sweeping method provided the best results for swimming pools. Conclusions Both sweeping methods are useful tools in epidemiological surveillance programs for the control of Aedes aegypti.

Organization of the cysts in bee (Hymenoptera, Apidae) testis: number of spermatozoa per cyst

The morphology of the cyst cells in Apis mellifera Linné, 1758, Scaptotrigona postica Latreille, 1804, and Melipona bicolor bicolor Lepeletier, 1836 testis, as well as the average number of spermatic cells are reported. The data indicates a supporting and nourrishing role of the cyst cells to the developing cystocytes. The counts of immature spermatozoa in the cysts show an average of 202.8 ± 21.2 spermatozoa for A. mellifera, 117.4 ± 8.68 for S. postica and 88.8 ± 15.57 for M. bicolor, which predict the occurrence of 8 mitotic cycles in the cystocytes of A. mellifera and 7 in the meliponines, considering that only one spermatozoom originates of each final spermatogonium.

Behavior of triatomines (Hemiptera, Reduviidae) vectors of Chagas' disease. II. Influence of feeding, lighting and time of day on the number of mating, mating speed and duration of copulation of Panstrongylus megistus (Burm, 1835) under laboratory conditions

To determine in influence of feeding, lighting and time of day on the copulating behavior of Panstrongylus megistus, 480 insect pairs were divided into four groups of 120 each and tested in the following respective situations: without food deprivation (F.D.), with five days of F.D., with ten days of F.D., and with 20 days of F. D. The tests were performed between 9:00 a.m. to 12:00a.m. and 7:00 p.m. to 10:00 p.m., with light (700-1400 lux) and in the dark (1.4-2.8 lux) and behavior was recorded by the time sampling technique. Mating spped (MS) and duration of copulation (DC) were also calculated for each situation. The maximum frequency of copulation was observed after five days of F.D., at night, in the dark (n = 16), and the minimum was observed for recently-fed pairs, at night, with light (n = 4). Males approached females more often than females approached males. MS was lowest in pairs with twenty days of F.D., at night, with light (X = 23.0 ± 16.0 minutes), and highest in recently-fed pairs, during the day, with light (X = 2.9 ± 2.5 minutes). DC was shortest in recently-fed insects, during the day, in the dark (X = 23.5 ± 6.7 minutes), and longest in recently-fed animals, at night, in the dark (X = 38.3 ± 6.9 minutes).

Behavior of triatomines (Hemiptera: Reduviidae) vectors of Chagas' disease: III. Influence of the number of matings on the fecundity and fertility of Panstrongylus megistus (Burm. 1835) in the laboratory

A study of the effect of mating in the fecundity and fertility of females of P. megistus fed on pigeon blood every 14 days, was carried out in the laboratory. Two groups were constituted: I - females which mated only once; II - females which stayed always with the males. Only 56.7% of group I females laid fertile eggs, while as much as 90% of group II females laid fertile eggs. The duration of the fertile oviposition was greater in the females which stayed always with the males. Some females of this group were able to mate up to seven times throughout their life-span. This fact render useless sterile males in the control of these insects. It is suggested that the components of pigeon's blood used for feeding the triatomines could have an influence upon the fecundity and fertility of the female sof the two groups.

Genetical exchanges between one Biomphalaria glabrata (Gastropoda: Planorbidae) and a varying number of partners

When one pigmented Biomphalaria glabrata is mated with 1 to 20 albino snails, the percentage of albino parent producing pigmented offspring decreases while the percentage of parent laying albino offspring increases. If the number of snaisl/group increases, the mean duration of the use of allosperm decreases.