Viremic blood donor found by a rapid screening method in a season of high human parvovirus B19 activity in Niterói, Rio de Janeiro, Brazil

Erythrovirus B19 infection is usually benign but may have serious consequences in patients with hemolytic anemia (transient aplastic crisis), immunodeficiency (in whom persistent infection can lead to chronic bone marrow failure with anemia), or who are in the first or second trimester of gestation (spontaneous abortion, hydrops fetalis, and fetal death). Being non-enveloped, B19 resists most inactivation methods and can be transmitted by transfusion. B19 is difficult to cultivate and native virus is usually obtained from viremic blood. As specific antibodies may be absent, and there is no reliable immunological method for antigen detection, hybridization or polymerase chain reaction are needed for detecting viremia. A rapid method, gel hemagglutination (Diamed ID-Parvovirus B19 Antigen Test), can disclose highly viremic donations, whose elimination lessens the viral burden in pooled blood products and may even render them non-infectious. In order to obtain native antigen and to determine the frequency of viremic donors, we applied this test to blood donors in a period of high viral activity in our community. Positive or indeterminate results were re-tested by dot-blot hybridization. We tested 472 donors in 1998 and 831 ones in 1999. One viremic donor was found in 1999. We suggest that in periods of high community viral activity the gel hemagglutination test may be useful in avoiding highly viremic blood being added to plasma pools or directly transfused to patients under risk.

Erythrovirus B19 infection in acquired immunodeficiency syndrome: screening by histopathology, immunohistochemistry, and in situ hybridization

Erythrovirus B19 infects erythrocytic progenitors, transiently interrupting erythropoiesis. In AIDS patients it causes chronic anemia amenable to treatment. We looked for evidences of B19 infection in stored bone marrow material from patients with acquired immunodeficiency syndrome. Histological sections were made from stored paraffin blocks from 33 autopsies (39 blocks) and 35 biopsies (45 blocks, 30 patients) performed from 1988 to 2002. They were examined after hematoxylin-eosin (HE) staining, immunohistochemical (IHC), and in situ hybridization. HE revealed intra-nuclear inclusion bodies ("lantern cells") suggesting B19 infection in 19 sections corresponding to 19 of 63 patients examined with this test. Seven of 78 sections subjected to immunohistochemistry were positive, corresponding to 7 of 58 patients examined with this test. Fourteen sections corresponding to 13 of the 20 HE and/or IHC positive patients were subjected to in situ hybridization, with six positives results. Among the 13 patients subjected to the three techniques, only one gave unequivocal positive results in all and was considered a true positive. The frequency of B19 infection (1/63 patients) in the material examined can be deemed low.

Screening of Amazonian plants from the Adolpho Ducke forest reserve, Manaus, state of Amazonas, Brazil, for antimicrobial activity

Tropical forests are species-rich reserves for the discovery and development of antimicrobial drugs. The aim of this work is to investigate the in vitro antimicrobial potential of Amazon plants found within the National Institute on Amazon Research's Adolpho Ducke forest reserve, located in Manaus, state of Amazonas, Brazil. 75 methanol, chloroform and water extracts representing 12 plant species were tested for antimicrobial activity towards strains of Mycobacterium smegmatis, Escherichia coli, Streptococcus sanguis, Streptococcus oralis, Staphylococcus aureus and Candida albicans using the gel-diffusion method. Active extracts were further evaluated to establish minimum inhibitory concentrations (MIC) and antimicrobial profiles using bioautography on normal-phase thin-layer chromatography plates. Diclinanona calycina presented extracts with good antimicrobial activity and S. oralis and M. smegmatis were the most sensitive bacteria. D. calycina and Lacmellea gracilis presented extracts with the lowest MIC (48.8 µg/ml). D. calycina methanol and chloroform leaf extracts presented the best overall antimicrobial activity. All test organisms were sensitive to D. calycina branch chloroform extract in the bioautography assay. This is the first evaluation of the biological activity of these plant species and significant in vitro antimicrobial activity was detected in extracts and components from two species, D. calycina and L. gracilis.

Evaluation of a malaria antibody enzyme immunoassay for use in blood screening

Transfusion-transmitted malaria is rare, but it may produce severe problem in the safety of blood transfusion due to the lack of reliable procedure to evaluate donors potentially exposed to malaria. Here, we evaluated a new enzyme-linked immunosorbent assay malaria antibody test (ELISA malaria antibody test, DiaMed, Switzerland) to detect antibodies to Plasmodium vivax (the indigenous malaria) in the blood samples in the Republic of Korea (ROK). Blood samples of four groups were obtained and analyzed; 100 samples from P.vivax infected patients, 35 from recovery patients, 366 from normal healthy individuals, and 325 from domestic travelers of non-endemic areas residents to risky areas of ROK. P.vivax antibody levels by ELISA were then compared to the results from microscopic examination and polymerase chain reaction (PCR) test. As a result, the ELISA malaria antibody test had a clinical sensitivity of 53.0% and a clinical specificity of 94.0% for P.vivax. Twenty out of 325 domestic travelers (6.2%) were reactive and 28 cases (8.6%) were doubtful. Of the reactive and doubtful cases, only two were confirmed as acute malaria by both microscopy and PCR test. Thus we found that the ELISA malaria antibody test was insufficiently sensitive for blood screening of P.vivax in ROK.

Synthesis of limonene β-amino alcohol derivatives in support of new antileishmanial therapies

A series of seven limonene β-amino alcohol derivatives has been regioselectively synthesised in moderate to good yields. Two of these compounds were found to be significantly effective against in vitro cultures of the Leishmania (Viannia) braziliensis promastigote form in the micromolar range. The activities found for 3b and 3f were about 100-fold more potent than the standard drug, Pentamidine, in the same test, while limonene did not display any activity. This is the first report of antileishmanial activity by limonene β-amino alcohol derivatives.

Risk factors associated with human papillomavirus infection in two populations from Rio de Janeiro, Brazil

We investigated human papillomavirus (HPV) infection in two female populations from diverse socio-economic strata from the state of Rio de Janeiro and we also investigated the possible co-factors related to infection and the progression to cancer. In Group I, the reference group of this study, 10.7% of the patients presented HPV infection, as detected by generic PCR, while in Group II (low socio-demographic conditions) HPV was detected in 31.1% of the samples. HPV16 was the most prevalent virus type found in both Groups I and II (5.3% and 10%, respectively), followed by HPV 18 (1.3% and 4.7%, respectively). Although only a small sample was analysed, we detected differences among the groups regarding the rates of HPV infection, HPV types, age, ethnicity, familial income, schooling, marital status, parity, tobacco smoking and oral contraceptive use. For Group I, the Papanicolaou test was the most powerful independent factor associated with HPV status, followed by an age of under 30 years old, the number of sexual partners and black ethnicity. Our data are in agreement with the co-factors that are typically described for the developed world. For Group II, the Pap test was also the most relevant variable that was analysed, but the history of other sexually transmitted diseases and the use of alcohol were additional factors that were implicated in infection. These findings point out the need for the development of general and specific strategies for HPV screening of all Brazilian women.

Malaria diagnosis from pooled blood samples: comparative analysis of real-time PCR, nested PCR and immunoassay as a platform for the molecular and serological diagnosis of malaria on a large-scale

Malaria diagnoses has traditionally been made using thick blood smears, but more sensitive and faster techniques are required to process large numbers of samples in clinical and epidemiological studies and in blood donor screening. Here, we evaluated molecular and serological tools to build a screening platform for pooled samples aimed at reducing both the time and the cost of these diagnoses. Positive and negative samples were analysed in individual and pooled experiments using real-time polymerase chain reaction (PCR), nested PCR and an immunochromatographic test. For the individual tests, 46/49 samples were positive by real-time PCR, 46/49 were positive by nested PCR and 32/46 were positive by immunochromatographic test. For the assays performed using pooled samples, 13/15 samples were positive by real-time PCR and nested PCR and 11/15 were positive by immunochromatographic test. These molecular methods demonstrated sensitivity and specificity for both the individual and pooled samples. Due to the advantages of the real-time PCR, such as the fast processing and the closed system, this method should be indicated as the first choice for use in large-scale diagnosis and the nested PCR should be used for species differentiation. However, additional field isolates should be tested to confirm the results achieved using cultured parasites and the serological test should only be adopted as a complementary method for malaria diagnosis.

Validation of a modified fluorimetric assay for the screening of trichomonacidal drugs

A fluorimetric microassay that uses a redox dye to determine the viability of the flagellate Trichomonas vaginalis has been optimised to provide a more sensitive method to evaluate potential trichomonacidal compounds. Resazurin has been used in recent years to test drugs against different parasites, including trichomonadid protozoa; however, the reproducibility of these resazurin-based methods in our laboratory has been limited because the flagellate culture medium spontaneously reduces the resazurin. The objective of this work was to refine the fluorimetric microassay method previously developed by other research groups to reduce the fluorescence background generated by the media and increase the sensitivity of the screening assay. The experimental conditions, time of incubation, resazurin concentration and media used in the microtitre plates were adjusted. Different drug sensitivity studies against T. vaginalis were developed using the 5-nitroimidazole reference drugs, new 5-nitroindazolinones and 5-nitroindazole synthetic derivatives. Haemocytometer count results were compared with the resazurin assay using a 10% solution of 3 mM resazurin dissolved in phosphate buffered saline with glucose (1 mg/mL). The fluorimetric assay and the haemocytometer counts resulted in similar percentages of trichomonacidal activity in all the experiments, demonstrating that the fluorimetric microtitre assay has the necessary accuracy for high-throughput screening of new drugs against T. vaginalis.

Applications of the hexanic fraction of Agave sisalana Perrine ex Engelm (Asparagaceae): control of inflammation and pain screening

The present study evaluated the anti-inflammatory and analgesic properties of Agave sisalana Perrine in classic models of inflammation and pain. The hexanic fraction of A. sisalana (HFAS) was obtained by acid hydrolysis followed by hexanic reflux. Anti-inflammatory properties were examined in three acute mouse models (xylene ear oedema, hind paw oedema and pleurisy) and a chronic mouse model (granuloma cotton pellet). The antinociceptive potential was evaluated in chemical (acetic-acid) and thermal (tail-flick and hot-plate test) models of pain. When given orally, HFAS (5, 10, 25 and 50 mg/kg) reduced ear oedema (p < 0.0001; 52%, 71%, 62% and 42%, respectively). HFAS also reduced hind paw oedema at doses of 10 mg/kg and 25 mg/kg (p < 0.05; 42% and 58%, respectively) and pleurisy at doses of 10 mg/kg and 25 mg/kg (41% and 50%, respectively). In a chronic model, HFAS reduced inflammation by 46% and 58% at doses of 10 mg/kg and 25 mg/kg, respectively. Moreover, this fraction showed analgesic properties against the abdominal writhing in an acetic acid model (at doses of 5-25 mg/kg) with inhibitory rates of 24%, 54% and 48%. The HFAS also showed an increased latency time in the hot-plate (23% and 28%) and tail-flick tests (61% and 66%) for the 25 mg/kg and 50 mg/kg doses, respectively. These results suggest that HFAS has anti-inflammatory and analgesic properties.

A simple and reliable method for the screening of transgenic tobacco plants

Even though much improvement has been made in plant transformation methods, the screening of transgenic plants is often a laborious work. Most approaches for detecting the transgene in transformed plants are still timeconsuming, and can be quite expensive. The objective of this study was to search for a simpler method to screen for transgenic plants. The infiltration of kanamycin (100 mg/mL) into tobacco leaves resulted in conspicuous chlorotic spots on the non-transgenic plant leaves, while no spots were seen on the leaves of transformed plants. This reaction occurred regardless of age of the tested plants, and the method has proven to be simple, fast, non-destructive, relatively cheap, and reliable. These results were comparable to those obtained by the polymerase chain reaction (PCR) amplification of the transgene using specific primers.

The role of fast atom bombardment mass spectroscopy (FABMS) in cluster characterization

Fast atom bombardment mass spectroscopy has been used to study a large number of cationic phosphine-containing transition-metal-gold clusters, which ranged in mass from 1000 to 4000. Many of these clusters have been previously characterized and were examined in order to test the usefulness of the FABMS technique. Results showed that FABMS is excellent in giving the correct molecular formula and when combined with NMR, IR, and microanalysis gave a reliable characterization for cationic clusters¹. Recently FABMS has become one of the techniques employed as routine in cluster characterization2,3 and also is an effective tool for the structure analysis of large biomolecules4. Some results in the present work reinforce the importance of these data in the characterization of clusters in the absence of crystals with quality for X-ray analysis.

Desenvolvimento de um spot test para o monitoramento da atividade da peroxidase em um procedimento de purificação

In this work we describe both a chromatographic purification procedure and a spot test for the enzyme peroxidase (POD: EC 1.11.1.7). The enzyme was obtained from crude extracts of sweet potatoes and the chromatographic enzyme purification procedure resulted in several fractions. Therefore a simple, fast and economic spot test for monitoring peroxidase during the purification procedure was developed. The spot test is based on the reaction of hydrogen peroxide and guaiacol, which is catalyzed by the presence of peroxidase yielding the colored tetraguaiacol.

Terbinafine: optimization of a LC method for quantitative analysis in pharmaceutical formulations and its application for a tablet dissolution test

A simple liquid chromatographic method was optimized for the quantitative determination of terbinafine in pharmaceutical hydroalcoholic solutions and tablets, and was also employed for a tablet dissolution test. The analysis was carried out using a RP-C18 (250 mm × 4.6 mm, 5 μm) Vertical® column, UV-Vis detection at 254 nm, and a methanol-water (95:5, v/v) mobile phase at a flow-rate of 1.2 mL min-1. Method validation investigated parameters such as linearity, precision, accuracy, robustness and specificity, which gave results within the acceptable range. The tablets dissolution was quite fast: 80% of the drug was dissolved within 15 min.

Análise exploratória rápida e não destrutiva (screening) da presença de elementos químicos tóxicos em material escolar por fluorescência de raios x

Compulsory in Brazil, toy certification aims to prevent possible risks in toy use. The Brazilian National Institute of Metrology (INMETRO) establishes the maximum concentration of toxic elements (TE) that may be present in toys. This study evaluates the presence of TE in different crayons, gouache and modeling clays using X-ray fluorescence. This technique is fast, has low operating cost and minimum sample pretreatment, resulting in a clean procedure without reagent consumption and waste generation. Bromine (in gouache) and Barium (in crayons) were the only potentially TE identified in the samples studied.

Use of alcohol before and after bariatric surgery

OBJECTIVE: to assess alcohol intake in the bariatric surgery pre and postoperative periods. METHODS: Patients were interviewed atSurgery Clinic of the Hospital das Clínicas da Universidade Federal de Pernambuco - HC/UFPE (Brazil) from July 2011 to March 2012. We analyzed socioeconomic, anthropometric and clinical variables. We used the Alcohol Use Disorders Identification Test (AUDIT C). RESULTS: One hundred nineteen patients were enrolled (mean age: 41.23+11.30 years), with a predominance of the female gender (83.2%), non-Caucasian race (55%), married individuals or in a stable union (65.5%), with a high school education (40.3%)and active in the job market (37%). Weight and body mass index (BMI) were 128.77+25.28Kg and 49.09+9.26Kg/m2,respectively in the preoperative period (class II obesity) and 87.19+19.16Kg and 33.04+6.21Kg/m2, respectively in the postoperative period (class I obesity) (p<0.001). Hypertension was the most frequent disease in the pre (66.6%) and postoperative (36.5%) periods. The prevalence of alcohol use was 26.6% in the preoperative period, of which 2.2% of high risk, and 35.1% in the postoperative period, of which 1.4% of probable dependence; this difference did not achieve statistical significance (p=0.337). CONCLUSION: The prevalence of abusive alcohol intake and/or probable dependence was low in both the pre and postoperative periods, with little evidence of risky consumption among the patients submitted to bariatric surgery.