On-line preconcentration system using a microcolumn packed with Alizarin Red S-modified alumina for zinc determination by flame atomic absorption spectrometry

A simple and sensitive on-line flow injection system for determination of zinc with FAAS has been described. The method is based on the separation and preconcentration of zinc on a microcolumn of immobilized Alizarin Red S on alumina. The adsorbed analyte is then eluted with 250 µL of nitric acid (1 mol L-1) and is transported to flame atomic absorption spectrometer for quantification. The effect of pH, sample and eluent flow rates and presence of various cations and anions on the retention of zinc was investigated. The sorption of zinc was quantitative in the pH range of 5.5-8.5. For a sample volume of 25 mL an enrichment factor of 144 and a detection limit (3S) of 0.2 µg L-1 was obtained. The precision (RSD, n=7) was 3.0% at the 20 µg L-1 level. The developed system was successfully applied to the determination of zinc in water samples, hair, urine and saliva.

Mathematical equation correction to spectral and transport interferences in high-resolution continuum source flame atomic absorption spectrometry: determination of lead in phosphoric acid

In this work, a new mathematical equation correction approach for overcoming spectral and transport interferences was proposed. The proposal was applied to eliminate spectral interference caused by PO molecules at the 217.0005 nm Pb line, and the transport interference caused by variations in phosphoric acid concentrations. Correction may be necessary at 217.0005 nm to account for the contribution of PO, since Atotal217.0005 nm = A Pb217.0005 nm + A PO217.0005 nm. This may be easily done by measuring other PO wavelengths (e.g. 217.0458 nm) and calculating the relative contribution of PO absorbance (A PO) to the total absorbance (Atotal) at 217.0005 nm: A Pb217.0005 nm = Atotal217.0005 nm - A PO217.0005 nm = Atotal217.0005 nm - k (A PO217.0458 nm). The correction factor k is calculated from slopes of calibration curves built up for phosphorous (P) standard solutions measured at 217.0005 and 217.0458 nm, i.e. k = (slope217.0005 nm/slope217.0458 nm). For wavelength integrated absorbance of 3 pixels, sample aspiration rate of 5.0 ml min-1, analytical curves in the 0.1 - 1.0 mg L-1 Pb range with linearity better than 0.9990 were consistently obtained. Calibration curves for P at 217.0005 and 217.0458 nm with linearity better than 0.998 were obtained. Relative standard deviations (RSD) of measurements (n = 12) in the range of 1.4 - 4.3% and 2.0 - 6.0% without and with mathematical equation correction approach were obtained respectively. The limit of detection calculated to analytical line at 217.0005 nm was 10 µg L-1 Pb. Recoveries for Pb spikes were in the 97.5 - 100% and 105 - 230% intervals with and without mathematical equation correction approach, respectively.

Soil chiseling and fertilizer location in sugarcane ratoon cultivation

While the pre-harvest sugarcane burning is a disused practice, green harvest requires changes concerning ratoon cultivation due to the presence of a thick layer of straw. The experiment, conducted in a mechanical green harvesting area cultivated with sugarcane, consisted of two stages: in the first stage, the mechanical straw cutting performance of flat disks with different geometry edges was evaluated, considering two types of disks and 10 replications in a completely randomized design; in the second stage, the effect of soil chiseling on both sides of planting lines, using shanks with straw cutting flat disks, was assessed, as well as fertilizer deposition form. The experimental design in the second stage was completely randomized, with seven treatments and five replications. Treatments consisted of a combination of two straw cutting disks (smooth or toothed edge), chiseling presence or absence, and fertilizer deposition forms (broadcast, on the planting line, and incorporated into chiseling furrows). The toothed disk differed from the smooth one, presenting lower values of horizontal and vertical forces, and torque. The agroindustrial variables pol (%), brix (%), fiber (%), and ATR (kg Mg-1) were not influenced by the fertilizer deposition form and soil chiseling. However, the localized fertilizer deposition increased crop yield when compared with broadcast fertilization.

Gene expression profile of ABC transporters and cytotoxic effect of ibuprofen and acetaminophen in an epithelial ovarian cancer cell line in vitro

PURPOSES: To determine the basic expression of ABC transporters in an epithelial ovarian cancer cell line, and to investigate whether low concentrations of acetaminophen and ibuprofen inhibited the growth of this cell line in vitro. METHODS: TOV-21 G cells were exposed to different concentrations of acetaminophen (1.5 to 15 μg/mL) and ibuprofen (2.0 to 20 μg/mL) for 24 to 48 hours. The cellular growth was assessed using a cell viability assay. Cellular morphology was determined by fluorescence microscopy. The gene expression profile of ABC transporters was determined by assessing a panel including 42 genes of the ABC transporter superfamily. RESULTS: We observed a significant decrease in TOV-21 G cell growth after exposure to 15 μg/mL of acetaminophen for 24 (p=0.02) and 48 hours (p=0.01), or to 20 μg/mL of ibuprofen for 48 hours (p=0.04). Assessing the morphology of TOV-21 G cells did not reveal evidence of extensive apoptosis. TOV-21 G cells had a reduced expression of the genes ABCA1, ABCC3, ABCC4, ABCD3, ABCD4 and ABCE1 within the ABC transporter superfamily. CONCLUSIONS: This study provides in vitro evidence of inhibitory effects of growth in therapeutic concentrations of acetaminophen and ibuprofen on TOV-21 G cells. Additionally, TOV-21 G cells presented a reduced expression of the ABCA1, ABCC3, ABCC4, ABCD3, ABCD4 and ABCE1 transporters.

Development and evaluation of a loop-mediated isothermal amplification (LAMP) assay for rapid detection of Actinobacillus pleuropneumoniae based the dsbE-like gene

This paper reports on the development and validation of a loop-mediated isothermal amplification assay (LAMP) for the rapid and specific detection of Actinobacillus pleuropneumoniae (A. pleuropneumoniae). A set of six primers were designed derived from the dsbE-like gene of A.pleuropneumoniae and validate the assay using 9 A. pleuropneumoniae reference/field strains, 132 clinical isolates and 9 other pathogens. The results indicated that positive reactions were confirmed for all A. pleuropneumoniae strains and specimens by LAMP at 63ºC for 60 min and no cross-reactivity were observed from other non-A.pleuropneumoniae including Haemophilus parasuis, Escherichia coli, Pasteurella multocida, Bordetella bronchiseptica, Streptococcus suis, Salmonella enterica, Staphylococcus, porcine reproductive and respiratory syndrome virus (PRRSV), and Pseudorabies virus. The detection limit of the conventional PCR was 10² CFU per PCR test tube, while that of the LAMP was 5 copies per tube. Therefore, the sensitivity of LAMP was higher than that of PCR. Moreover, the LAMP assay provided a rapid yet simple test of A. pleuropneumoniae suitable for laboratory diagnosis and pen-side detection due to ease of operation and the requirement of only a regular water bath or heat block for the reaction.

Cyanobacterial occurrence and detection of microcystin by planar chromatography in surface water of Billings and Guarapiranga Reservoirs, SP, Brazil

Billings and Guarapiranga Reservoirs were deeply affected by environmental disturbances, which more evident consequence are the cyanobacterial blooms. Microcystins are the most common cyanotoxin in freshwaters and more than 70 types are known. Different methods for microcystins analysis in water can be used, among which ELISA and HPLC are the most frequently employed. However, less sophisticated and more economic methods can also be used. This is the case of planar chromatography (thin-layer chromatography) method previously used in cyanotoxins purification but gradually replaced by others. Posterior optimization of the microcystin chromatography conditions and because of its simplicity, rapidity, efficiency and low cost, this method is again considered an option for the analysis of microcystins and nodularins. Considering the importance of Billings and Guarapiranga Reservoirs for drinking water supplies and the few scientific data about cyanobacteria and cyanotoxins in these water bodies, the aims of this work are to analyze the biodiversity of cyanobacteria in the Billings and Guarapiranga Reservoirs and the detection of dissolved microcystins in the water. It was possible to identify 17 species of cyanobacteria, 9 of them being potentially toxic. In Billings Reservoir Microcystis aeruginosa (Kützing) Kützing and Cylindrospermopsis raciborskii (Woloszynska) Seenayya & Subba Raju are the most common species, while in Guarapiranga Reservoir only M. aeruginosa was considered as a common species. Microcystins were detected in all Billings Reservoir samples and in only one sample from Guarapiranga Reservoir.

IgM and IgA Antibody Responses in 12 Cases of Human Acquired Toxoplasmosis

The persistence, in some subjects, of specific IgM antibodies to Toxoplasma gondii for several months after the acute phase of infection has complicated the interpretation of serological test results for toxoplasmosis. Several reports have emphasized the value of the detection of Toxoplasma-specific IgA antibodies for the diagnosis of acute toxoplasmosis. In this article, we report the follow-up profiles of Toxoplasma-specific IgM and IgA antibodies in serum samples obtained from 12 patients at various intervals after the onset of the clinical manifestations of infection. IgM antibodies were detected by the indirect immunofluorescence (IIF) test, antibody capture enzyme-linked immunosorbent assay (cELISA) and enzyme-mediated chemilluminescent technique (CmL). IgA antibodies were quantified by the direct ELISA (dELISA) and cELISA procedures. As defined by the manufacturer of the cELISA test for IgA used, most patients with acute toxoplasmosis have antibody levels > 40 arbritary units per ml (AU/ml). At values > 40 AU/ml, the cELISA for IgA detected significant antibody levels for a shorter time than the other techniques used for IgM and IgA detection. However, IgA levels £ 40 AU/ml do not exclude the possibility of acute toxoplasmosis since such levels can be reached very soon after infection with T. gondii. The results obtained in the present study show that the serological diagnosis of acute toxoplasmosis may not be such an easy task. Our data suggest that use of the IgA-cELISA concomitantly with IgM antibody screening could permit, in some circumstances, a more efficient diagnosis of acute acquired toxoplasmosis

Comparison of serology, antigenemia assay and the polymerase chain reaction for monitoring active cytomegalovirus infections in hematopoietic stem cell transplantation patients

Forty-six allogeneic hematopoietic stem cell transplantation (HSCT) patients were monitored for the presence of CMV antibodies, CMV-DNA and CMV antigens after transplantation. Immunoenzymatic serological tests were used to detect IgM and the increase in CMV IgG antibodies (increase IgG), a nested polymerase chain reaction (N-PCR) was used to detect CMV-DNA, and an antigenemia assay (AGM) was used to detect CMV antigens. The presence of CMV-IgM and/or CMV-increase IgG antibodies was detected in 12/46 (26.1%) patients, with a median time between HSCT and the detection of positive serology of 81.5 days. A positive AGM was detected in 24/46 (52.2%) patients, with a median time between HSCT and antigen detection of 62 days. Two or more consecutive positive N-PCR results were detected in 32/46 (69.5%) patients, with a median time between HSCT and the first positive PCR of 50.5 days. These results confirmed that AGM and mainly PCR are superior to serology for the early diagnosis of CMV infection. Six patients had CMV-IgM and/or CMV-increase IgG with a negative AGM (five cases) or N-PCR assay (one case). In five of these cases the serological markers were detected during the first 100 days after HSCT, the period of highest risk. These findings support the idea that serology may be useful for monitoring CMV infections in HSCT patients, especially when PCR is unavailable.

NEW PRIMERS FOR DETECTION OF Leishmania infantum USING POLYMERASE CHAIN REACTION

SUMMARY Leishmania infantum causes visceral leishmaniasis (VL) in the New World. The diagnosis of VL is confirmed by parasitological and serological tests, which are not always sensitive or specific. Our aim was to design new primers to perform a Polymerase Chain Reaction (PCR) for detecting L. infantum. Sequences of the minicircle kinetoplast DNA (kDNA) were obtained from GenBank, and the FLC2/RLC2 primers were designed. Samples of DNA from L. infantum, Leishmania amazonensis, Leishmania braziliensis, Leishmania guyanensis, Leishmania naiffi, Leishmania lainsoni, Leishmania panamensis, Leishmania major and Trypanosoma cruzi were used to standardize the PCR. PCR with FLC2/RLC2 primers amplified a fragment of 230 bp and the detection limit was 0.2 fg of L. infantum DNA. Of the parasite species assayed, only L. infantum DNA was amplified. After sequencing, the fragment was aligned to GenBank sequences, and showed (99%) homology with L. infantum. In the analysis of blood samples and lesion biopsy from a dog clinically suspected to have VL, the PCR detected DNA from L. infantum. In biopsy lesions from humans and dogs with cutaneous leishmaniasis, the PCR was negative. The PCR with FLC2/RLC2 primers showed high sensitivity and specificity, and constitutes a promising technique for the diagnosis of VL.

An optimized one-tube, semi-nested PCR assay for Paracoccidioides brasiliensis detection

Introduction Herein, we report a one-tube, semi-nested-polymerase chain reaction (OTsn-PCR) assay for the detection of Paracoccidioides brasiliensis. Methods We developed the OTsn-PCR assay for the detection of P. brasiliensis in clinical specimens and compared it with other PCR methods. Results The OTsn-PCR assay was positive for all clinical samples, and the detection limit was better or equivalent to the other nested or semi-nested PCR methods for P. brasiliensis detection. Conclusions The OTsn-PCR assay described in this paper has a detection limit similar to other reactions for the molecular detection of P. brasiliensis, but this approach is faster and less prone to contamination than other conventional nested or semi-nested PCR assays.

Influenza infection and Kawasaki disease

INTRODUCTION: The objective of this study was to investigate the possible link between influenza (Flu) infection and Kawasaki disease (KD). METHODS: We examined the medical records of 1,053 KD cases and 4,669 influenza infection cases hospitalized at our institute from January 1, 2011 to December 31, 2013. Cases of KD with concomitant influenza infection formed the KD + Flu group. Each KD + Flu case was matched with 2 KD cases and 2 influenza infection cases, and these cases were assigned to the KD group and Flu group, respectively. The differences in the principal clinical manifestations, course of disease, incomplete KD rate, intravenous immunoglobulin (IVIG) resistance rate, and echocardiographic detection results between the KD + Flu group and KD group were compared. The fever durations and laboratory test results of these three groups were compared. RESULTS: 1) The seasonal variations of the KD + Flu group, KD group and Flu group were similar. 2) The morbidity rate of incomplete KD was higher in the KD + Flu group compared with the KD group. 3) Patients in the KD + Flu group exhibited a longer time to KD diagnosis compared with patients in the KD group. 4) The KD + Flu group exhibited the longest fever duration among the three groups. 5) The CRP and ESR values in the KD + Flu group were higher those in the Flu or KD groups. CONCLUSIONS: Concomitant influenza infection affects the clinical manifestations of KD and can impact the laboratory test results and the diagnosis and treatment of the disease. However, it remains unclear whether influenza contributes to KD etiology.

Doses and methods of distribution of potassium chloride on soybean crop (Glycine max (L.) Merrill) in a red latosol, affecting soil salinity, grain production and chemical composition of leaves

The present work deal t wi th an experiment under field conditions and a laboratory test of soil incubation the objectives were as follows: a. to study effects on soybean grain product ion and leaf composition of increasing doses of potassium chloride applied into the soil through two methods of distribution; b. to observe chemical modifications in the soils incubated with increasing doses of potassium chloride; and, c. to correlate field effects with chemical alterations observed in the incubation test, The field experiment was carried out in a Red Latosol (Haplustox) with soybean cultivar UFV - 1. Potassium chloride was distributed through two methods: banded (5 cm below and 5 cm aside of the seed line) and broadcasted and plowed-down. Doses used were: 0; 50; 100 and 200 kg/ha of K2O. Foliar samples were taken at flowering stage. Incubation test were made in plastic bags with 2 kg of air dried fine soil, taken from the arable layer of the field experiment, with the following doses of KC1 p,a. : 0; 50; 100; 200; 400; 800; 1,600; 3.200; 6,400 and 12,800 kg/ha of K(2)0. In the conditions observed during the present work, results allowed the following conclusions: A response by soybean grain production for doses of potassium chloride, applied in both ways, banded or broadcasted, was not observed. Leaf analysis did not show treatment influence over the leaf contents for N, P, K, Ca, Mg, and CI, Potassium chloride salinity effects in both methods of distribution for all the tested closes were not observed.

Soil invertebrates in southern Brazilian Araucaria forest - grassland mosaic: differences between disturbed and undisturbed areas

Soil invertebrate distribution in Araucaria forest, grassland and edge habitats was studied in both disturbed and undisturbed areas in southern Brazil. Mean-density and taxa compositions were verified. Invertebrate densities differed between grassland and the other two habitats in the undisturbed area but not across the disturbed one. At the disturbed area taxa differed between the grassland and the other two habitats. The undisturbed area, on the other hand, presented taxa differences only between the grassland and the forest habitats. Acari, Arachnida and Collembola were the most sensitive taxa for detecting differences across habitats in both areas. At the disturbed area, these taxa presented densities lowering from the forest to the grassland. At the undisturbed area the same taxa increased from the forest to the grassland. Coleoptera and Formicidae (Insecta) presented no difference between habitats at the studied taxonomic level.

Morphological development of Corydoras aff. paleatus (Siluriformes, Callichthyidae) and correlation with the emergence of motor and social behaviors

Here we examine major anatomical characteristics of Corydoras aff. paleatus (Jenyns, 1842) post-hatching development, in parallel with its neurobehavioral evolution. Eleutheroembryonic phase, 4.3-8.8 days post-fertilization (dpf); 4.3-6.4 mm standard length (SL) encompasses from hatching to transition to exogenous feeding. Protopterygiolarval phase (8.9-10.9 dpf; 6.5-6.7 mm SL) goes from feeding transition to the commencement of unpaired fin differentiation, which marks the start of pterygiolarval phase (11-33 dpf; 6.8-10.7 mm SL) defined by appearance of lepidotrichia in the dorsal part of the median finfold. This phase ends with the full detachment and differentiation of unpaired fins, events signaling the commencement of the juvenile period (34-60 dpf; 10.8-18.0 mm SL). Eleutheroembryonic phase focuses on hiding and differentiation of mechanosensory, chemosensory and central neural systems, crucial for supplying the larval period with efficient escape and nutrient detection-capture neurocircuits. Protopterygiolarval priorities include visual development and respiratory, digestive and hydrodynamic efficiencies. Pterygiolarval priorities change towards higher swimming efficacy, including carangiform and vertical swimming, necessary for the high social interaction typical of this species. At the end of the protopterygiolarval phase, simple resting and foraging aggregations are seen. Resting and foraging shoals grow in complexity and participant number during pterygiolarval phase, but particularly during juvenile period.

Analysis of the interacting components between larval Schistosoma mansoni and schistosome-susceptible and resistant Biomphalaria glabrata

Cell-free hemolymph (serum) and hemocytes from Schistosoma mansoni-susceptible (PR albino M-line) and resistant (10-R2) strains of Biomphalaria glabrata were compared by SDS-PAGE, immunoblotting and radioiodination. Whole serum of both snail strains is dominated by hemoglobin (Hb) (MW = 160 Kd). SDS-PAGE. of Hb-depleted serum indicated that the 10-R2 strain has dominant polypeptides in the 50 to 30 Kd range whereas PR albino snails have few low MW proteins. Antibodies raised to whole PR albino and 10-R2 serum, and the 160 Kd (Hb) band reacted similarly in immunoblot assays. Analysis of hemocytes revealed that 10-R2 snails have a surface-exposed protein at about 80 Kd which is not present on PR albino hemocytes. An examination of primary cultured sporocysts indicated the presence of four major surface proteins (40, 50, 55, 70 Kd) and two minor surface-exposed polypeptides (92, 170 Kd). Antibodies raised against live, intact sporocysts reacted almost exclusively with sporocyst-surface proteins when tested by immunoblotting.