Tratamento de ferida em Coendou prehensilis (Rodentia: Erethizontidae) com laserterapia e ozonioterapia: relato de caso / Wound treatment of Coendou prehensilis (Rodentia: Erethizontidae) with laser and ozone therapy: case report

O emprego conjunto da laserterapia e da ozonioterapia em feridas apresenta alto potencial benéfico para os pacientes, uma vez que contribui para o manejo da dor, tem ação anti-inflamatória e acelera o processo de cicatrização. Este relato de caso tem como objetivo apresentar o uso de terapias alternativas na cicatrização de ferida em exemplar de Coendou prehensilis. Um ouriço-cacheiro, fêmea, adulto, com peso de 4kg foi encaminhado para atendimento médico veterinário com histórico de ter sido atacado por um cão. Inicialmente o ouriço passou pelo procedimento de higienização e desbridamento da ferida, para a retirada das bordas necróticas. Adicionalmente, foram administrados clindamicina (10mg/kg), por via intramuscular (IM), uma vez por dia (SID), tramadol (4mg/kg, IM, SID), flunixin (0,3mg/kg, SID), por via subcutânea (SC), e ferrodextrano (25mg/kg, IM, SID). Apesar da terapia instituída, observou-se reincidência de crescimento necrótico tecidual, o que levou à eleição do tratamento da ferida com as técnicas de laserterapia e ozonioterapia. O emprego das terapias alternativas como adjuvante promoveu uma cicatrização satisfatória da ferida, com ausência de sinais de sensibilidade local e de infecção, bem como ausência de crescimento de bordas necróticas. O tratamento adjuvante foi eficaz e pode ser empregado em outras situações para cicatrização de ferida em mamíferos silvestres.(AU)

The use of therapy with laser beam and ozone in wounds has a high beneficial potential for patients, since it contributes to the management of pain, has an anti-inflammatory action and accelerates the cicatricial process. Due to this casuistry importance, the case report aims to present alternative therapy use for wound healing on a Coendou prehensilis. Thus, a female of C. prehensilis weighing 4kg, was sent to veterinary care. At first there was a hygiene process and debridement for necrotic edge removal. Furthermore, injected clindamycin (10mg/kg) was administered intramuscularly (IM), once a day (SID), tramadol (4mg/kg, IM, SID), flunixin (0.3mg/kg, SID), administered subcutaneously (SC) and iron dextran (25mg/kg, IM, SID). In spite of the established therapy, tissue necrotic growth was observed, which lead the wound treatment as healing by second intention, initiating an alternative therapy with laser beam and ozone. As a result, the healing was satisfactory due to the elected techniques, without signs of pain and infection. The adjuvant treatment with physiotherapy had advantageous effect and could be applied to wound healing in wild mammal animals.(AU)

Antimutagenicity and antigenotoxicity of Aloe arborescens Miller and Aloe barbadensis Miller in Aspergillus nidulans and Wistar rats.

Medicinal plants such as Aloe arborescens Miller and Aloe barbadensis Miller are used by the general population to treat various diseases. Therefore, the aim of this study was to evaluate the antimutagenicity of these two species using a methG1 system in Aspergillus nidulans and the comet assay in rats. The animals were treated with the plants at concentrations of 360 and 720 mg/kg body weight (1 and 2, respectively) by gavage for 14 days, followed by the administration of etoposide on treatment day 8. Blood samples were prepared for analysis of DNA damage. For the test in A. nidulans, the biA1methG1 lineage conidia were treated for 4 h with both plant species at concentrations of 4 and 8% (w/v). Then, they were washed and plated on a selective medium for frequency analysis of survival and mutation. The results of the comet assay showed that both plants were antigenotoxic compared to etoposide, which was not a typical response of methG1 systems, where only the highest concentration of plant extracts usually exhibit beneficial effects. This study demonstrates the potential antigenotoxicity and antimutagenicity of the Aloe plants tested and, therefore, supports their use as a form of preventive therapy and for health maintenance by the population.

Composição centesimal, compostos bioativos e atividade antioxidante de frutos de Morinda citrifolia Linn (noni) cultivados no Paraná / Proximate composition, bioactive compounds and antioxidant activity of fruits of Morinda citrifolia L. (noni) cultivated in Paraná, Brazil

O fruto de noni (Morinda citrifolia L.) é consumido há milênios na medicina popular polinésia devido aos benefícios nutricionais e terapêuticos. O consumo de noni em outros países, incluindo o Brasil, cresceu vertiginosamente nos últimos anos em decorrência das atividades biológicas atribuídas a ingestão do suco da fruta, principalmente pela propriedade anticâncer. Contudo, a composição química da planta, que está relacionada com suas propriedades biológicas, é determinada pelo seu local de origem, e por influência do clima e do solo onde é cultivada. Neste sentido, este trabalho teve como objetivo analisar a polpa extraída de frutos maduros de noni cultivados em Maringá-PR. A análise da polpa in natura apresentou 89,16% de umidade, 0,75% de cinzas, 2,10% de proteínas, 2,19% de lipídios e 5,81% de carboidratos. Dos compostos bioativos, foram analisados antocianinas (1,39 mg.100 g-1 polpa), flavonoides amarelos (13,01 mg.100 g-1 polpa), carotenóides (0,45 mg.100 g-1 polpa) e vitamina C (12,16 mg.100 g-1 polpa). Para fenólicos totais e atividade antioxidante foram preparados diferentes extratos, sendo que os maiores teores de fenólicos totais foram encontrados no extrato aquoso (1143,56 mg equivalente de ácido gálico (EAG).100 g-1), seguido do extrato etanólico (966,96 mg EAG.100 g-1), metanol/acetona (820,88 mg EAG.100 g-1) e metanólico (306,33 mg EAG.100 g-1). Os melhores resultados para antioxidantes, determinado pelo EC50 - concentração do extrato necessária para reduzir 50% do radical DPPH, foram encontrados nos extratos metanol/acetona (EC50 de 25,18 mg.mL-1) e metanólico (EC50 de 25,96 mg.mL-1). A atividade antioxidante dos frutos pode estar relacionada com o conteúdo de vitamina C, uma vez que os extratos com um menor conteúdo de fenóis totais foram aqueles que apresentaram menores valores de EC50.

The noni fruit (Morinda citrifolia L.) has been consumed for millennia in the Polynesian folk medicine because of its nutritional and therapeutic benefits. The consumption of the noni fruit in other countries, including Brazil, has increased in recent years because of the biological activities attributed to the ingestion of noni juice, especially the anticancer effect. However, the chemical composition of the plant, which is related to its biological properties, is determined by its geographical origin and is influenced by climate and soil. Therefore, this study aimed to analyze the pulp extracted from noni fruit grown in Maringá, state of Paraná, Brazil. The fresh pulp analysis showed 89.16% of moisture, 0.75% of ash, 2.10% of protein, 2.19% of fat and 5.81% of carbohydrates. The bioactive compounds analyzed were anthocyanins (1.39 mg.100 g-1 pulp), yellow flavonoids (13.01 mg.100 g-1 pulp), carotenoids (0.45 mg.100 g-1 pulp) and vitamin C (12.16 mg.100 g-1 pulp). For the analysis of total phenols and antioxidant activity, different extracts were prepared. The highest total phenolic contents were found in the aqueous extract (1143.56 mg EAG.100 g-1), followed by the ethanol extract (966.96 mg EAG.100 g-1), methanol/acetone extract (820.88 mg EAG.100 g-1) and methanol extract (306.33 mg EAG.100 g-1). The results for the antioxidant capacity were determined by the EC50, which is concentration of extract required to reduce 50% of the DPPH radical. The best results for the antioxidant capacity were found in the methanol/acetone extract (EC50 of 25.18 mg.mL-1) and in the methanol extract (EC50 of 25.96 mg.mL-1). The antioxidant activity of the fruit may be related to its vitamin C content, since the extracts with lower total phenolic content were those that had lower EC50 values.

Efeito da Aloe arborescens Miller e da Aloe barbadensis Miller sobre o desenvolvimento vegetativo em Aspergillus nidulans / Effects of the Aloe arborescens Millerand Aloe barbadensis Milleron over vegetative growth in the Aspergillus nidulans

RESUMOA pesquisa de produtos naturais benéficos à saúde humana vem crescendo nos últimos 20 anos. Considerando que as plantas de Aloe são amplamente utilizadas pela população humana, em geral de maneira terapêutica, o objetivo deste estudo foi avaliar os efeitos de Aloearborescens Miller e Aloe barbadensis Miller, sobre o desenvolvimento vegetativo de linhagens normais e mutantes de Aspergillus nidulans. Conídios da linhagem biA1methG1, MSE e CLB3 de A. nidulans, foram inoculados em meio completo sem (Controle) e com extratos das duas espécies incubados por 2, 4, 6 e 8 horas a 37ºC, no escuro. Foi analisado em microscópio óptico, 200 conídios de cada tratamento. Para o desenvolvimento das colônias, as linhagens foram inoculadas no centro das placas juntamente com o meio de cultura sólido e sobre a membrana de diálise, visando a medição do diâmetro e do peso. A análise estatística foi baseada no teste de Tukey e todos os procedimentos experimentais foram conduzidos em triplicata. Todas as linhagens apresentaram interferências positivas quando expostas às plantas de Aloe, porém, de maneira variada. Ambas as espécies aceleraram a germinação em todas as linhagens testadas e atuaram na redução significativa de conídios mortos e/ou malformados. Em relação ao desenvolvimento vegetativo, todos os dados referentes ao peso úmido e diâmetro corrigido dos tratamentos demonstraram progressos, contudo, a razão diâmetro/peso apresentou somente na linhagem MSE, ação favorável dos tratamentos naturais. As informações deste estudo sugerem benefícios de A. arborescens e A. barbadensis, justificando a importância e continuidade da investigação, para melhor elucidar os mecanismos de ação dessas plantas.

ABSTRACTThe researches about natural products that arebeneficial to human health have been growing over the past 20 years. Since Aloe plants are broadly used by the general population, frequently due to therapeutic reasons, the objective of this study was to evaluate the effects of Aloe arborescens Millerand Aloe barbadensis Miller on the vegetative growth of normal and mutant strains of Aspergillus nidulans. The conidia of thebiA1methG1, MSE and CLB3 strains of A. nidulans were inoculated in complete environment without (control) and with extracts of two species of Aloeincubated for 2, 4, 6 and 8 hours at 37˚C. 200 conidia were analyzed by optical microscopy. For the development of the colonies, the strains were inoculated in the center of the plates together with the solid environment of the cultivation and over the dialysis membrane for measuring the diameter and weighing. The statistical analysis was based on the Tukey test and all experimental procedures were performed in triplicate. All strains showed positive interference when exposed to Aloe plants, however, through different manners. Both species have accelerated the germination in all tested strains and acted in the significant reduction of dead and / or malformed conidia. Regarding the vegetative growth, all data related to wet weight and corrected diameter of the treatments revealed progress, however, the ratio diameter/weightpresented improvement only in the MSE lineage, favorable action of natural treatments. The information from this study suggest that A. arborescens and A. barbadensis are beneficial, thus justifying the importance of research maintenance in order to better elucidate the action mechanisms of these plants.

Morphoanatomic characterization of the stem and the leaf of Tabernaemontana catharinensis A. DC (Apocynaceae) and antimutagenic activity of its leaves / Caracterização morfoanatômica do caule e da folha de Tabernaemontana catharinensis A. DC (Apocynaceae) e atividade antimutagênica de suas folhas

ABSTRACT Tabernaemontana catharinensis A. DC (Apocynaceae) is used as a medicinal plant by the population. In order to contribute to the safe use of the plant as herbal medicine, this study aimed to morphoanatomically characterize the aereal vegetative organs of T. catharinensis and to evaluate the leaves’ mutagenic and antimutagenic activities. Histological blades of leaves and stem of T. catharinensis were performed; the methionine system (methG1) and Aspergillusnidulans conidia germination analysis were employed for mutagenic and antimutagenic evaluation. The morphoanatomic analysis did not show trichomes in the stem, petiole and leaf. Besides, it was observed both the presence of bi-collateral bundles - except in the foliar apex where the bundles were from the collateral type - as well as anamphistomatic leaf with paracyte stomata and sub-epidermal layer in the region of the leaf edges. The mutagenicity/antimutagenicity trial indicated a significant decrease of mutation frequency in comparison with the control group and showed that the T. catharinensis had antimutagenic activity within the type, time and form of treatment. Since the germination test showed that the conidia germination was accelerated from the bud phase, activities at the cell cycle level and polarized growth proved to be possible. The morphoanatomic analysis of the leaf and stem associated with the mutagenic and antimutagenic analyses contributes to the safe use of the plant by humans and also for the quality control of a possible phytotherapeutic drug.

RESUMO Tabernaemontana catharinensis A. DC (Apocynaceae) é utilizada como planta medicinal pela população. A fim de contribuir para o uso seguro da planta como medicinal, este trabalho teve como objetivo caracterizar morfoanatomicamente os órgãos vegetativos aéreos de T. catharinensis e avaliar a atividade mutagênica e antimutagênica de suas folhas. Foram realizados cortes histológicos da folha e do caule de T. catharinensis e, para a avaliação mutagênica e antimutagênica, foi utilizado o sistema metionina (methG1) e análise da germinação de conídios em Aspergillus nidulans. A análise morfoanatômica evidenciou a ausência de tricomas no caule, pecíolo e folha; presença de feixes bicolaterais, com exceção no ápice foliar cujos feixes são do tipo colateral; folha anfiestomática com estômatos paracíticos e camada subepidérmica na região do bordo foliar. O ensaio de mutagenicidade/antimutagenicidade mostrou uma diminuição significativa da frequência de mutação em relação ao controle, indicando que nesse tipo, tempo e forma de tratamento, T. catharinensis apresentou atividade antimutagênica. O ensaio de germinação evidenciou que houve aceleração da germinação dos conídios, a partir da fase de botão, indicando uma possível atuação em nível de ativação de ciclo celular e crescimento polarizado. A análise morfoanatômica da folha e do caule associados à análise mutagênica e antimutagênica, contribuem para o uso seguro da planta pela população e para o controle de qualidade de um possível fitoterápico.

Trans-resveratrol concentrations and antimutagenic potential of juice from the grape cultivars Vênus, BRS Violeta and Isabel.

Grape juice, in addition to being an energetic food, due to its high sugar content, has several compounds that can prevent or treat various types of diseases. Resveratrol is a compound present in grapes that has attracted a lot of interest; in addition to preventing cardiovascular disease linked to lipid metabolism, it has chemopreventive and chemotherapeutic activities. We evaluated the antimutagenic activity and determined the trans-resveratrol content in grape juice from the varieties Vênus, BRS Violeta and Isabel. The grape juices from the three cultivars and the resveratrol solution were tested in the methG1 system in Aspergillus nidulans. The conidia from the biA1methG1 strain were treated for 4 h in 10% grape juice (v/v). After washing, the conidia were placed in selective media to analyze survival and mutations. The standard resveratrol solution and the grape juice of the cultivar Isabel, both with a trans-resveratrol content of 1 mg/mL, presented antimutagenic potential in this test system because the frequency of mutation of the treatments was significantly lower than the frequency of spontaneous mutation. However, grape juice from the varieties Vênus and BRS Violeta, both with a lower quantity of trans-resveratrol, gave weak antimutagenic activity in this test system because the frequency of mutation of the treatments was significantly higher than the frequency of spontaneous mutation.

Toxicity and genotoxicity of water and sediment from streams on dotted duckweed (Landoltia punctata).

Most rivers are used as a source to supply entire cities; the quality of water is directly related to the quality of tributaries. Unfortunately men have neglected the importance of streams, which receive domestic and industrial effluents and transport nutrients and pesticides from rural areas. Given the complexity of the mixtures discharged into these water bodies, this study aimed to evaluate the quality of water and sediment of ten tributaries of Pirapó River, in Maringá, Paraná State, Brazil. To this end, the free-floating macrophyte Landoltia punctata (G. Meyer) Les & D.J.Crawford was used as test organism in microcosm, and the toxicity of water and sediment samples was evaluated by the relative growth rate, dry/fresh biomass ratio, and genotoxic effects (comet assay). Samples of water and sediment of each stream were arranged in microcosms with L. punctata. Seven days later, plants were collected for analysis. Nutrient levels were higher than the reference location, indicating eutrophication, but the results indicated a toxic effect for only three streams, and a genotoxic effect for all streams.

In situ analysis of apoptosis in Aspergillus nidulans with ethidium bromide and acridine orange.

Apoptosis and necrosis are among several types of cell death. We stained the nuclei of Aspergillus nidulans grown in micro-colonies with ethidium bromide and acridine orange to detect in situ apoptosis. Suspensions of conidia from 5-day-old colonies of the A. nidulans strains biA1methG1, G422, CLC100, and CLB3 were each put into two tubes. The suspension of one tube was irradiated with ultraviolet light for 20 s, whereas the other tube was not exposed to irradiation. The two suspensions were inoculated in complete liquid medium and 50-µL samples were placed on sterilized cover slips, spread on the surface of solid culture media on Petri dishes. After the micro-colonies were formed, the material on the cover slips was stained with ethidium bromide and acridine orange, placed on the lamina and observed under a fluorescence microscope. This staining method was efficient in discriminating normal nuclei from those going apoptosis and necrosis. Results have shown that irradiation provokes apoptosis but does not induce necrosis. There were no differences between the three strains and all data were considered to be statistically significant.

Avaliação da mutagenicidade de Piper methysticum L. f. no sistema methG1 em Aspergillus nidulans / Mutagenic evaluation of Piper methysticum L. f. in Aspergillus nidulans methG1 system

O extrato seco da raiz de Piper methysticum L. f. Forster (PIPERACEAE), a kava-kava, é usado no tratamento de diversos problemas envolvendo ansiedade como um dos sintomas. Por não causar dependência, sedação e ter ação ansiolítica, muitas pessoas têm recorrido a kava-kava para auxiliá-las no emagrecimento. Isto pode levar ao consumo indiscriminado da planta e acarretar riscos, pois todo medicamento fitoterápico deve respeitar limites de doses. Um risco na utilização de plantas medicinais é a toxicidade e, dentro deste, a mutagenicidade. Como a mutagenicidade está relacionada com a carcinogenicidade torna-se importante testar este potencial na kava-kava. Assim, o objetivo deste trabalho foi avaliar o potencial mutagênico do extrato seco da raiz de P. methysticum no sistema methG1 em Aspergillus nidulans. A linhagem utilizada foi a biA1methG1, auxotrófica para biotina e metionina. Conídios dormentes de colônias crescidas por cinco dias foram tratados com soluções da kava-kava nas concentrações de 0,35 mg mL-1 e 3,5 mg mL-1, e depois de 24h, semeados em meio seletivo contendo metionina, para análise dos sobreviventes, e sem metionina, para a análise dos mutantes. Os números de sobreviventes e mutantes dos tratamentos foram comparados aos do controle. Os resultados indicaram que o extrato da raiz da kava-kava é mutagênica, pois a freqüência de mutação dos tratamentos foi maior que da mutação espontânea, porém não ocorrendo diferença significativa entre as doses.

The dry root extract of Piper methysticum L. f. Forster (Piperaceae), kava-kava, is used as to treat several health problems involving anxiety symptoms. As it causes no addiction, it can be applied as a sedative and anxiolytic. Many people have been relying on kava-kava as an auxiliary treatment. This can lead to an indiscriminate plant consumption and lead to risks, because all phytotherapic medications must observe dosage limits. One risk in the folk medicinal plant use is toxicity, and within it, mutagenicity. As mutagenicity is closely related to carcinogenicity, it is important to test the kava-kava mutagenicity potential. Thus, the purpose of this work was to test the mutagenicity of the dry root extract of P. methysticum in the methG1 system of Aspergillus nidulans. The bia1methG1 lineage, which is auxotrophic for biotine and methione, was used. Conidia from five-day-old colonies were collected and treated with kava-kava solutions at 0.35 mg mL-1 and 3.5 mg mL-1 concentrations and, after 24h, they were planted in selective growth medium with and without methione, in order to analyze the survivors and mutants, respectively. The number of survivors and mutants analyzed under effect of the treatments was compared with the control. The results indicated that the kava-kava dry root extract is mutagenic, since the mutation frequency of the treatments was higher than spontaneous mutation, however, there were no differences between the doses tested.

Morphometric and quantitative analysis of theafferent renal artery variation

The kidney is a retroperitoneal organ that weight from 125 to 170 g in the adult men and 115to 155 g in adult women. Irrigation kidney is characterized by the presence of large anatomical variability thatmay be influenced by ethnic and to a lesser extent by gender. Among the variations, there may be the presenceof an accessory renal artery that is projected into the upper or lower end of the kidney. This research aimsto observe the incidence of anatomical variations of the afferent renal artery and quantify both right and leftkidney weight.Materials and Methods:We analyzed kidney weights and accessory renal artery variations in48 adult kidneys of both genders obtained from Institute of Anatomy of the University of Severino Sombra.Subsequently, we compared the mean weights of kidneys in order to ascertain whether there was significantdiscrepancy between the left and the right kidney. For this, we performed the Student t test considering aP < 0.05.Results:The mean weight of the right kidney was 140.4 ± 22.6 g and the left was 148.8 ± 20.5 g.In 40% of right kidneys was observed anatomic variation with the presence of accessory renal artery. To the leftkidney was observed a variation of 35%.Conclusion:We found that the accessory renal artery, when present,was more closely related to the end of the kidney especially in the right kidney.

PCR analyses of tRNA intergenic spacer, 16S-23S internal transcribed spacer, and randomly amplified polymorphic DNA reveal inter- and intraspecific relationships of Enterobacter cloacae strains.

PCR analysis of tRNA intergenic spacer (tDNA-PCR) and of the 16S-23S internal transcribed spacer (ITS-PCR) and random amplified polymorphic DNA (RAPD) analysis were evaluated for their usefulness in characterization of Enterobacter cloacae strains isolated from both clinical origins and vaccine microbial contamination. tDNA-PCR presented specific and reproducible patterns for Enterobacter sakazakii ATCC 29004, Enterobacter aerogenes ATCC 13048, and Enterobacter cloacae ATCC 13047 and 23355 that presented the same profile for all 16 E. cloacae isolates, offering an alternative tool for species-level identification. ITS-PCR and RAPD analysis yielded completely different banding patterns for the 20 strains studied, except for E. cloacae strains isolated from different batches of vaccine that exhibited a unique pattern, suggesting contamination by the same strain. The combined use of tDNA-PCR and ITS-PCR in a one-step protocol allows accurate identification and typing of E. cloacae strains a few hours after the colony has been isolated.

Isotopic evidence for variations in the marine calcium cycle over the Cenozoic.

Significant variations in the isotopic composition of marine calcium have occurred over the last 80 million years. These variations reflect deviations in the balance between inputs of calcium to the ocean from weathering and outputs due to carbonate sedimentation, processes that are important in controlling the concentration of carbon dioxide in the atmosphere and, hence, global climate. The calcium isotopic ratio of paleo-seawater is an indicator of past changes in atmospheric carbon dioxide when coupled with determinations of paleo-pH.

Identification and characterization of a novel fibronectin-binding protein on the surface of group A streptococci.

Understanding the role surface proteins play in the interaction of group A streptococci with epithelial cells is an important step toward the development of new strategies to fight infections. Fibronectin-binding proteins in streptococci and staphylococci have been described as important mediators for adherence to eukaryotic cells. In the present study we describe a new Streptococcus pyogenes fibronectin-binding protein (PFBP). The gene encoding the PFBP protein (pfbp) was identified from an M12 strain genomic library. It encodes a protein of 127.4 kDa which contains the LPXTGX motif characteristic of cell wall-associated proteins in gram-positive organisms and is among the largest surface molecules described for group A streptococci. The pfbp gene is transcribed during cell growth and was present in several class I and II streptococcal strains tested. The deduced amino acid sequence of PFBP exhibits a variable N-terminal region and a conserved C-terminal region when compared to most fibronectin-binding proteins identified from other gram-positive bacteria. The N-terminal region presents a stretch of 105 amino acids with no homology with N-terminal regions of previously described fibronectin-binding molecules, while the C-terminal region contains three repeat domains that share significant similarity with the repeat regions of fibronectin-binding proteins from S. pyogenes, S. dysgalactiae, and S. equisimilis. The PFBP repeated region, when expressed on the surface of S. gordonii, a commensal organism, binds to soluble and immobilized fibronectin. This study also shows that, in addition to pfbp, a second gene homologous with that of protein F1 (which also codes for a fibronectin-binding protein) is transcribed during cell growth in the same S. pyogenes strain.

RS-102221: a novel high affinity and selective, 5-HT2C receptor antagonist.

The 5-HT2C receptor is one of three closely related receptor subtypes in the 5-HT2 receptor family. 5-HT2A and 5-HT2B selective antagonists have been described. However, no 5-HT2C selective antagonists have yet been disclosed. As part of an effort to further explore the function of 5-HT2C receptors, we have developed a selective 5-HT2C receptor antagonist, RS-102221 (a benzenesulfonamide of 8-[5-(5-amino-2,4-dimethoxyphenyl) 5-oxopentyl]-1,3,8-triazaspiro[4.5]decane-2,4-dione). This compound exhibited nanomolar affinity for human (pKi = 8.4) and rat (pKi = 8.5) 5-HT2C receptors. The compound also demonstrated nearly 100-fold selectivity for the 5-HT2C receptor as compared to the 5-HT2A and 5-HT2B receptors. RS-102221 acted as an antagonist in a cell-based microphysiometry functional assay (pA2 = 8.1) and had no detectable intrinsic efficacy. Consistent with its action as a 5-HT2C receptor antagonist, daily dosing with RS-102221 (2 mg/kg intraperitoneal) increased food-intake and weight-gain in rats. Surprisingly, RS-102221 failed to reverse the hypolocomotion induced by the 5-HT2 receptor agonist 1-(3-chlorophenyl)piperazine (m-CPP). It is concluded that RS-102221 is the first selective, high affinity 5-HT2C receptor antagonist to be described.

Identification and characterization of a new protein from Streptococcus pyogenes having homology with fibronectin and fibrinogen binding proteins.

Fibronectin and fibrinogen-binding proteins have been described as possible adhesin in streptococci and staphylococci. Recent published data has demonstrated that Protein F, a fibronectin-binding protein from group A streptococci, is important in adherence to respiratory cells (8). Other similar proteins already described (i.e. SOF, Sfb and SfbII) are able to competitively inhibit the binding of fibronectin to S. pyogenes (9,10,5). Similarly, clumping factor from S. aureus, is known to promote adherence to fibrinogen-coated surfaces (7). When the sequence from SFFBP-12 was compared against all the others fibronectin and fibrinogen-binding proteins described in streptococci and staphylococci (1-10), an identity at the amino acid level, ranging from 38 to 69% was found for the C region. For the repeated region (R), the identity ranged between 47 and 75%. Unlike all the other proteins already described in group A streptococci, the protein we describe here, SFFBP-12, shares a high degree homology (67-75%) with the fibronectin-binding protein B from S. dysgalactiae (6), as well as homology with the S. aureus clumping factor (7) and fibronectin-binding protein B (3), making it a new potential fibronectin-fibrinogen binding protein for group A streptococci. These characteristics would also imply that SFFBP-12 contains two different fibronectin-binding domains (regions B and C), thus enhancing its role as a possible major adhesin molecule. RNA transcription assays showed a transcript with the expected molecular size for the intact SFFBP-12 protein, confirming that the protein is actively expressed during bacterial growth. SFFBP-12 is the largest protein of its kind identified from group A streptococci and is comparable in size to the fibronectin binding protein B from S. dysgalactiae (6). If it is shown that SFFBP-12 does in fact bind both fibronectin and fibrinogen, as the sequence data suggest, it would make this molecule a major virulence determinant for the group A streptococcus.

Purification, recovery, and laser-driven fluorination of silicon from dissolved and particulate silica for the measurement of natural stable isotope abundances.

A procedure for the purification, recovery, and determination of isotopic abundances of silicon from biogenic and lithogenic particulate matter and dissolved silicic acid is reported. Purification involves the reaction of acid molybdate with dissolved silicon in natural waters or that produced by the dissolution of particulate silica by hydrofluoric acid. The resulting silicomolybdic acid is then quantitatively precipitated by reaction with triethylamine hydrochloride. The silicon is recovered as silicon dioxide through stepwise combustion of the dried precipitate. Fluorination of the product for isotopic analysis is accomplished by laser heating under pure fluorine generated by the decomposition of a fluorine-based salt. The resulting silicon tetrafluoride is separated from hydrogen fluoride and other fluorination byproducts cryogenically using a variable-temperature cold trap. Yields for silicon recovery are 99.9% for precipitation and greater than 95% for the purification/fluorination procedure. Reproducibility of the isotopic composition for pure quartz granules processed through the procedure is ±0.1‰ for δ(30)Si.

A species-specific nucleotide sequence of Mycobacterium tuberculosis encodes a protein that exhibits hemolytic activity when expressed in Escherichia coli.

Species-specific proteins may be implicated in the unique pathogenic mechanisms characteristic of Mycobacterium tuberculosis. In previous studies, a 3.0-kb species-specific DNA fragment of M. tuberculosis was identified (C. A. Parra, L. P. Londoño, P. del Portillo, and M. E. Patarroyo, Immun. 59:3411-3417, 1991). The nucleotide sequence of this 3.0-kb fragment has been obtained. This sequence was shown to contain two open reading frames (ORFs) whose putative gene products share 68.9% identity between each other. The major ORF shows 57.8% similarity with PLC-N and 53.2% similarity with PLC-H, two phospholipase C enzymes from Pseudomonas aeruginosa. The major ORF was amplified by PCR and cloned into the pGEX-5T expression vector. Cell extracts of Escherichia coli overexpressing this glutathione S-transferase fusion protein were shown to produce beta-hemolysis suggestive of phospholipase activity. Since phospholipase C enzymes have been reported as virulence factors of P. aeruginosa and also of the intracellular pathogen Listeria monocytogenes, it is possible that the proteins identified in this study could also play a role in sustaining tuberculosis infection in humans.

Vaccination with SPf66, a chemically synthesised vaccine, against Plasmodium falciparum malaria in Colombia.

Preclinical and clinical studies have established the safety and immunogenicity of the chemically synthesised SPf66 malaria vaccine. The present study is a phase III randomised, double-blind, placebo-controlled, efficacy trial completed in La Tola, Colombia. 1548 volunteers over one year of age received three doses of either the vaccine (n = 738) or placebo (n = 810). Active and passive case detection methods were used to document clinical episodes of malaria among the study population. The follow-up period began one month after the third dose and lasted for one year. 168 and 297 episodes of Plasmodium falciparum malaria were documented in the SPf66 group and the placebo group, respectively; this corresponds to a crude protective efficacy of 38.8%. Incidence rates for first or only P falciparum malarial episodes were 22.3% per annum among the vaccinee group and 33.5% among the placebo group (RR = 1.5; 95% Cl 1.23, 1.84). Therefore, the protective efficacy of SPf66 against first or only episodes was 33.6% (95% Cl 18.8, 45.7), being highest in children aged 1-4 years (77%) and adults older than 45 years (67%). The estimated protective efficacy against second episodes was 50.5% (95% Cl 12.9-71.9). Our study shows that the chemically synthesised SPf66 malaria vaccine is safe, immunogenic, and protective against P falciparum malaria in semi-immune populations subject to natural challenge.

Safety and immunogenicity of the synthetic malaria vaccine SPf66 in a large field trial.

In the first field trial with synthetic malaria vaccine SPf66 in a large population naturally exposed to malaria, 9957 persons greater than 1 year old and residing on the Colombian Pacific coast received three doses of the vaccine. To evaluate vaccine safety, clinical observations were made 30 min and 48 h after each immunization. There were no adverse reactions in 95.7% of cases. In the 4.3% of cases with adverse reactions, local induration and erythema were the most frequent. In a randomly selected group of vaccinees, anti-SPf66 antibody titers were measured after the third dose: 93% of the vaccinees raised antibodies to SPf66. Among these, 55% had titers greater than 1:1600. These results demonstrate the safety and immunogenicity of the SPf66 vaccine in a large field trial.