A determinação direta do nitrogênio nítrico, amoniacal e amídico (uréia) em fertilizantes

O presente trabalho tem por objetivo o estudo da determinação direta de diversas formas de nitrogênio (nítrico, amoniacal e amídico) em fertilizantes simples e em misturas. Constatada a presença das três formas citadas de nitrogênio, o extrato aquoso de 250 ml, obtido pelo tratamento de 2,5000 g da amostra, pode ser destinado às mencionadas determinações. Uma alíquota de 50 ml é transferida para uma coluna de resina de troca aniônica DOWEX-1-X8, que, convenientemente preparada, retém o ânion nitrato(NO-3), separando-o de outras formas. Em seguida, determina-se o nitrogênio nítrico na solução obtida pela eluição da coluna com solução de NaCl a 20%. O nitrogênio amoniacal pode ser determinado em uma alíquota do extrato aquoso original ou na solução que passa através da coluna de resina para a separação do nitrato. O nitrogênio amídico (uréia) é determinado numa alíquota do extrato original pelo método da urease. Os métodos e técnicas preconizados foram préviamente estudados, através de determinações em soluções contendo substâncias puras (nitrato de sódio p.a., sulfato de amônio p.a. e uréia p.a.). Numa segunda fase, procedeu-se a aplicação dos mencionados métodos e técnicas na determinação do nitrogênio em misturas de fertilizantes preparadas em laboratório, em fertilizantes comerciais simples e em misturas também comerciais. Os resultados obtidos indicam que os métodos e técnicas recomendados permitem a determinação direta e relativamente rápida do nitrogênio nítrico, amoniacal e amídico (uréia) em fertilizantes.

Influencia do leite puro ou fermentado sobre a curva do peso

A ministração a coelhos, pela via digestiva, de leite fresco - crú ou fervido, e de leite fermentado, quer naturalmente, quer artificialmente, pela proliferação de bacilos coli ou aerogenes, não influe na curva de peso de maneira bastante apreciavel. A maior perda de peso, observada num coelho que ingeria leite fresco, não fervido, poude ser attribuida a modificações da flora do intestino e suas consequencias sobre o organismo do animal. Estes resultados estão de accôrdo com observações feitas durante o preparo de sôros immunisantes anti-coli em coelhos, aos quaes propinavamos quantidades consideraveis de corpos bacterianos, sem que lhes occorresse perda significativa de peso.

Acção do leite-crú ou fervido, e do leite fermentado, sobre o intestino perfundido segundo o methodo de Cohnheim - Magnus

Leite fresco-crú ou fervido nenhuma acção exerce sobre o tonus de uma preparação de intestino isolado e perfundido, entretanto determina augmento bastante apreciavel da amplitude das contracções rhythmicas. O leite fermentado, natural ou experimentalmente, pela proliferação de bacilos coli ou aerogenes, por espaço de 6 a 24 horas, em estufa a 37° C., determina muitas vezes augmento consideravel do tonus do segmento de intestino, bem assim augmento da amplitude das oscilações phasicas, em todo caso, uma e outra alteração mostrando-se de modo menos energico do que já observaramos com os filtrados de culturas antigas de bacillo coli. A acção sobre o tonus não parece depender da acidez do leite fermentado, desde que ella não desapparece com a neutralisação do leite. De outro lado, uma solução de acído lactíco do mesmo theôr encontrado no leite fermentado, expresso em graus Dornic, não determina augmento do tonus da preparação. O aquecimento até 115° C., ou o resfriamento prolongado a + 3° C., não supprime a actividade do leite fermentado natural - ou artificialmente, no que concerne seus efeitos sobre tonus da preparação de intestino isolado, facto analogo tendo sido já por nós observado com os filtrados de culturas antigas de bacillo coli. A ingestão prévia de filtrados de culturas antigas de bacillo coli, ou de leite fermentado experimentalmente por esse germe, parece determinar uma maior reactividade do segmento de intestino, o que se póde averiguar pelas reacções mais promptas e mais energicas, quando dos ensaios in-vitro com leite fermentado experimentalmente com bacillo coli, em confronto com as experiencias realisadas com intestino de coelhos normaes. O mesmo facto já havia sido por nós observado com os filtrados de culturas antigas de bacillo coli.

Sôbre u'a modificação do meio de Monteverde

It is well known that the culture media used in the presumptive diagnosis of suspiciuous colonies from plates inoculated with stools for isolation of enteric organisms do not always correctly indicate the major groups of enterobacteria. In an effort to obtain a medium affording more exact indications, several media (1-9) have been tested. Modifications of some of these media have also been tested with the result that a satisfactory modification of Monteverde's medium was finaly selected. This proved to be most satisfactory, affording, as a result of only one inoculation, a complete series of basic indications. The modification involves changes in the formula, in the method of preparation and in the manner of storage. The formulae are: A. Thymol blue indicator: NaOH 0.1/N .............. 34.4 ml; Thymol blue .............. 1.6 g; Water .................... 65.6 ml. B. Andrade's indicator. C. Urea and sugar solution: Urea ..................... 20 g; Lactose ................... 30 g; Sucrose ................... 30 g; Water .................... 100 ml. The mixture (C.) should be warmed slightly in order to dissolve the ingredients rapidly. Sterilise by filtration (Seitz). Keep stock in refrigeratior. The modification of Monteverde's medium is prepared in two parts. Semi-solid part - Peptone (Difco) 2.0 g; NaCl 0.5 g; Agar 0.5 g; Water 100.0 ml. Boil to dissolve the ingredients. Adjust pH with NaOH to 7.3-7.4. Boil again for precipitation. Filter through cotton. Ad indicators "A" 0.3 ml and "B" 1.0 ml. Sterilise in autoclave 115ºC, 15 minutes in amounts not higher than 200 ml. Just before using, add solution "C" asseptically in amounts of 10 ml to 200 ml of the melted semi-solid medium, maintained at 48-50ºC. Solid part - Peptone (Difco) 1.5 g; Trypticase (BBL) 0.5 g; Agar 2.0 g; Water 100,00 ml. Boil to dissolve the ingredients. Adjust pH with NaOH to 7.3-7.4. Boils again. Filter through cotton. Add indicators "A" 0.3 ml and "B" 1.0 ml; ferrous ammonium sulfate 0.02 g; sodiun thiosulfate 0.02 g. Sterilise in autoclave 115ºC, 15 minutes in amounts not higher than 200 ml. Just before using, add solution "C" asseptically in amounts of 10 ml to 200 ml of the melted solid medium, maintained at 48-50ºC. Final medium - The semi-solid part is dispensed first (tubes about 12 x 120 mm) in 2.5 ml amounts and left to harden at room temperature, in vertical position. The solid part is dispensed over the hardened semi-solid one in amounts from 2.0 ml to 2.5 ml and left to harden in slant position, affording a butt of 12 to 15 mm. The tubes of medium should be subjected to a sterility test in the incubator, overnight. Tubes showing spontaneous gas bubbles (air) should then be discarded. The medium should be stored in the incubator (37ºC), for not more than 2 to 4 days. Storage of the tubes in the ice-box produces the absorption of air which is released as bubbles when the tubes are incubated at 37ºC after inoculation. This fact confirmed the observation of ARCHAMBAULT & McCRADY (10) who worked with liquid media and the aplication of their observation was found to be essential to the proper working conditions of this double-layer medium. Inoculation - The inoculation is made by means of a long straight needle, as is usually done on the triple sugar, but the needel should penetrate only to about half of the height of the semi-solid column. Indol detection - After inoculation, a strip of sterelized filter papaer previously moistened with Ehrlich's reagent, is suspended above the surface of the medium, being held between the cotton plug and the tube. Indications given - In addition to providing a mass of organisms on the slant for serological invetigations, the medium gives the following indications: 1. Acid from lactose and/or sucrose (red, of yellowsh with strains which reduce the indicators). 2. Gas from lactose and/or sucrose (bubbles). 3. H[2]S production, observed on the solid part (black). 4. Motility observed on the semi-solid part (tubidity). 5. Urease production, observed on solid and semi-solid parts (blue). 6. Indol production, observed on the strip of filter paper (red or purplish). Indol production is not observed with indol positive strains which rapidly acidify the surface o the slant, and the use of oxalic acid has proved to give less sensitive reaction (11). Reading of results - In most cases overnight incubation is enough; sometimes the reactions appear within only a few hours of incubation, affording a definitive orientation of the diagnosis. With some cultures it is necessary to observe the medium during 48 hours of incubation. A description showing typical differential reaction follows: Salmonella: Color of the medium unchanged, with blackening of the solid part when H[2]S is positive. The slant tends to alkalinity (greenish of bluish). Gas always absent. Indol negative. Motility positive or negative. Shigella: Color of the medium unchanged at the beginning of incubation period, but acquiring a red color when the strain is late lactose/sucrose positive. Slant tending to alkalinity (greenish or purplish). Indol positive or negative. Motility, gas and H[2]S always negative. Proteus: Color of the medium generally changes entirely to blue or sometimes to green (urease positive delayed), with blackening of solid part when H[2]S is positive. Motility positive of negative. Indol positive. Gas positive or negative. The strains which attack rapidly sucrose may give a yellow-greenish color to the medium. Sometimes the intense blue color of the medium renders difficult the reading of the H[2]S production. Escherichiae and Klebsiellae: Color of the medium red or yellow (acid) with great and rapid production of gas. Motility positive or negative. Indol generally impossible to observe. Paracoli: Those lactose of sucrose positive give the same reaction as Esherichia. Those lactose or sucrose negatives give the same reactions as Salmonellae. Sometimes indol positive and H[2]S negative. Pseudomonas: Color of the medium unchanged. The slant tends to alkalinity. It is impossible to observe motility because there is no growth in the bottom. Alkaligenes: Color of the medium unchanged. The slant tends to alkalinity. The medium does not alter the antigenic properties of the strains and with the mass of organisms on the slant we can make the serologic diagnosis. It is admitted that this medium is somewhat more laborious to prepare than others used for similar purposes. Nevertheless it can give informations generally obtained by two or three other media. Its use represents much saving in time, labor and material, and we suggest it for routine laboratory work in which a quick presumptive preliminary grouping of enteric organisms is needed.

The interaction of Gram negative bacteria and S. mansoni in mice with experimental Schistosomiasis

Animals (122 mice) were infected each with eighty cercariae of S. mansoni and subsequently challenged intravenously eight weeks later with the following gram-negative organisms. S. typhi, E. coli, Klebsiella-enterobacter species, Proteus mirabilis and Pseudomonas aeruginosa. Enumeration of bacteria in the liver, spleen and blood and S. mansoni from the portal sistem was performed from one to four weeks later in infected animals. A significant difference between infection produced by S. typhi and other gram negative organisms was observed: S. typhi persisted longer in the spleen and liver and could be recovered from S. mansoni worms up to three weeks following bacterial infection. Other gram negative bacteria disappeared from S. mansoni worms after two weeks of initial challenge. Additional animals (51 mice) infected with S. mansoni were given S. typhi, E. coli or sterile saline. After two weeks, animals were sacrificed and the recovery rate of worms from the portal system, and the mesenteric and hepatic oogram were determined. in animals infected with E. coli a significant decrease in the number of worms was observed compared to the saline control group; thirty worms were recovered in the control group compared to two worms in e. coli infected animals. In addition, the patterns of oviposition was significantly different in these latter animals suggesting complete inhibition of this process. Following S. typhi infection the difference in recovery of worms and pattern of oviposition was minimal. These findings suggest a difference in the interaction of various gram negative bacteria and S. mansoni and are consistent with the clinical observation of prolonged salmonella bacteremia in patients with schistosomiasis.

Bactérias gram negativas resistentes a antimicrobianos em alimentos

A partir de 154 espécimens de alimentos, representados por hortaliças (alface), leite e merenda escolar, obteve-se o isolamento e identificação de 400 amostras de bacilos Gram negativos. Esta amostragem se distribuiu em 339 enterobactérias (Escherichia, Shigella, Citrobacter, Klebsiella, Enterobacter, Serratia e Proteus) e 61 de gêneros afins (Acinetobacter, Flavobacterium, Aeromonas e Pseudomonas). Submetendo-se as culturas aos antimicrobianos: sulfadiazina (Su), estreptomicina (Sm), tetraciclina (Tc), cloranfenicol (Cm), canamicina (Km), ampicilina (Ap), ácido nalidíxico (Nal) e gentamicina (Gm), observou-se apenas seis estirpes sensíveis a todas as drogas e sensibilidade absoluta à Gm. A predominância dos modelos Su (27,6%) e Su-Ap (39,6%) incidiu nas enterobactérias, enquanto que, 18,0% para Ap e 9,8% para Su-Ap foram detectados nos gêneros afins. Para caracterização da resistência foram realizados testes de conjugação e a totalidade das culturas não revelou transferência para o gene que confere resistência ao ácido nalidíxico. Relevantes são as taxas de amostras R+ observadas nos bacilos entéricos, oscilando em torno de 90% (leite e merenda escolar) e alface, em torno de 70%

Immunomodulation by microbial ribosomes

Over the past twenty years, many authors have reported evidence of the immunoprotective capacity of ribosomes isolated from bacteria, fungi and parasites. Since 1971 we have explored the protective capacity of ribosomes isolated from a large variety of microorganisms responsible for human and animal diseases. More recently, using monoclonal antibodies raised against ribosomes and then selected for their ability to confer passive immunity to mice, we have studied the mechanism of the protection induced by ribosomes. These studies, in parallel with the development of a technology for the large scale production of ribosomes, have allowed us to achieve a new regard for ribosomal vaccines for use in human. The general concept of ribosomal vaccines in presented and examples of two such vaccines are described with data on the specific protection that they induce in mice against experimental infections with Klebsiella peneumoniae, Streptococcus pneumoniae, S. pyogenes and Haemophilus influenzae for the first one, and against Candida albicans type A and type B for the second one. Because of their high immunogenicity and their innocuity these vaccines represent a decisive improvement over classical microbial vaccines.

In vitro cell activating properties of the composite ribosomal vaccine D53

D53 (RibomuntyR) is a composite vaccine made of immunogenic ribosomes from 4 bacterial species (Klebsiella pneumoniae, Haemophilus influenzae, Streptococcus pyogenes and Streptococcus pneumoniae) associated with a membrane proteoglycan from a non encapsulated strain of Klebsiella pneumoniae. D53 is a potent inducer of interleukin-1 production by mouse BALB/c spleen cells as shown by the C3H/HeJ thymocyte co-stimulation assay. Furthermore D53 triggers DNA synthesis by mouse spleen cells and induces the maturation of B lymphocytes into immunoglobulin secreting cells. Polyclonal B cell activation by D53 was readily achieved in the C3H/HeJ strain which is deficient in its response to E. coli lipopolysaccharide. The proliferative response to D53 was abrogated by removal of B cells from the spleen cell suspension, but it was not altered after depletion of T cells or adherent cells. D53 induced polyclonal B cell activation of spleen cells from athymic nude mice and from CBA/N mice. Each component of D53 induced polyclona B cell activation except ribosomes from Streptococcus pneumoniae. Each triggered Interleukin-1 synthesis except ribosomes from Klebsiella penumoniae. These in vitro properties may account for some of the in vivo immunostimulating properties of this composite vaccine.

The sirtuin inhibitor cambinol impairs MAPK signaling, inhibits inflammatory and innate immune responses and protects from septic shock.

Sirtuins (SIRT1-7) are NAD(+)-dependent histone deacetylases (HDACs) that play an important role in the control of metabolism and proliferation and the development of age-associated diseases like oncologic, cardiovascular and neurodegenerative diseases. Cambinol was originally described as a compound inhibiting the activity of SIRT1 and SIRT2, with efficient anti-tumor activity in vivo. Here, we studied the effects of cambinol on microbial sensing by mouse and human immune cells and on host innate immune responses in vivo. Cambinol inhibited the expression of cytokines (TNF, IL-1β, IL-6, IL-12p40, and IFN-γ), NO and CD40 by macrophages, dendritic cells, splenocytes and whole blood stimulated with a broad range of microbial and inflammasome stimuli. Sirtinol, an inhibitor of SIRT1 and SIRT2 structurally related to cambinol, also decreased macrophage response to TLR stimulation. On the contrary, selective inhibitors of SIRT1 (EX-527 and CHIC-35) and SIRT2 (AGK2 and AK-7) used alone or in combination had no inhibitory effect, suggesting that cambinol and sirtinol act by targeting more than just SIRT1 and SIRT2. Cambinol and sirtinol at anti-inflammatory concentrations also did not inhibit SIRT6 activity in in vitro assay. At the molecular level, cambinol impaired stimulus-induced phosphorylation of MAPKs and upstream MEKs. Going well along with its powerful anti-inflammatory activity, cambinol reduced TNF blood levels and bacteremia and improved survival in preclinical models of endotoxic shock and septic shock. Altogether, our data suggest that pharmacological inhibitors of sirtuins structurally related to cambinol may be of clinical interest to treat inflammatory diseases.

Helicobacter pylori in dyspeptic children and adults: endoscopic, bacteriologic and histologic correlations

Using different bacteriological (urease test, Gram staining and culture) and histological (Steiner staining and modified Giemsa staining) techniques, we searched for the presence of Helicobacter pylori in the gastric antrum of 200 dyspeptic Brazilian patients (106 females and 94 males aged 19 days to 81 years). The presence of bacteria was then correlated with the endoscopic and histological findings. H. pylori was present in 59.5 of the population studied. In Brazil, colonization occurs early, involving 37 of the dyspeptic population by 20 years of age. The presence of H. pylori in the gastric antrum was strongly associated with duodenal ulcer (P < 0.001) and a normal endoscopic examination did not exclude the possibility of colonization of the gastric antrum by H. pylori. The most sensitive test was the preformed urease test (89). We conclude that more than one diagnostic method should preferably be used for the detection of H. pylori and that the presence of H. pylori is closely correlated with active chronic gastritis (P < 0.001).

Antibacterial activity of Ocimum gratissimum L. essential oil

The essential oil (EO) of Ocimum gratissimum inhibited Staphylococcus aureus at a concentration of 0.75 mg/ml. The minimal inhibitory concentrations (MICs) for Shigella flexineri, Salmonella enteritidis, Escherichia coli, Klebsiella sp., and Proteus mirabilis were at concentrations ranging from 3 to 12 mg/ml. The endpoint was not reached for Pseudomonas aeruginosa (>=24 mg/ml). The MICs of the reference drugs used in this study were similar to those presented in other reports. The minimum bactericidal concentration of EO was within a twofold dilution of the MIC for this organism. The compound that showed antibacterial activity in the EO of O. gratissimum was identified as eugenol and structural findings were further supported by gas chromatography/mass spectra retention time data. The structure was supported by spectroscopic methods.

Epidemiologia de la Peritonitis Bacteriana Espontània en el nostre medi

Objectius: . Descriure l’epidemiologia de les PBE del nostre hospital. Material i Mètodes: S’analitzen les característiques de les PBE diagnosticades entre 2007-2010. Resultats: S’identificaren 44 episodis (73% extrahospitalàries). La taxa de cultius positius fou del 52% (52%BGN; 34%CGP). S’observà : 18% d’Escherichia coli amb resistència a quinolones; 57% de Klebsiella spp BLEA. Aparegué insuficiència renal en un 20%. La mortalitat fou del 27%. Conclusions: No s’observà una major proporció de CGP en el grup nosocomial. La resistència a quinolones i la producció de BLEA és inferior a la reportada. La mortalitat fou superior en el grup nosocomial.

Non-tuberculous mycobacteria I: one year clinical isolates identification in Tertiary Hospital Aids Reference Center, Rio de Janeiro, Brazil, in pre highly active antiretroviral therapy era

The aim of this study was to determine the prevalence of non-tuberculous mycobacteria (NTM) isolates at University Hospital, Reference Center for Aids in Rio de Janeiro, Brazil, during one year. We used standard biochemical tests for species identification and IS1245 PCR amplification was applied as a Mycobacterium avium specific identification marker. Four hundred and four specimens from 233 patients yielded acid-fast bacilli growth. M. tuberculosis was identified in 85% of the patients and NTM in 15%. NTM disseminated infection was a common event correlated with human immunodeficiency virus (HIV) infected patients and only in HIV negative patients the source of NTM was non sterile site. M. avium complex (MAC) was biochemically identified in 57.8% (49/83) of NTM isolates, most of them from sterile sites (75.5%), and in 94% (46/49) the IS 1245 marker specific for M. avium was present. Twenty NTM strains showed a MAC biochemical pattern with the exception of a urease-positive (99% of MAC are urease-negative), however IS1245 was detected in 96% of the strains leading to their identification as M. avium. In this group differences in NTM source was not significant. The second most frequently isolated NTM was identified as M. scrofulaceum (7.2%), followed by M. terrae (3.6%), M. gordonae (2.4%), M. chelonae (1.2%), M. fortuitum (1.2%) and one strain which could not be identified. All were IS1245 negative except for one strain identified as M. scrofulaceum. It is interesting to note that non-sterile sites were the major source of these isolates (92.8%). Our finding indicated that M. avium is still the major atypical species among in the MAC isolates recovered from Brazilian Aids patients without highty active antiretroviral therapy schema. Some discrepancies were seen between the identification methods and further investigations must be done to better characterize NTM isolates using other phenotypic and genotypic methods.

Detection of pathogenic bacteria in skin lesions of patients with chiclero's ulcer: reluctant response to antimonial treatment

We investigated the bacterial flora present in skin lesions of patients with chiclero's ulcer from the Yucatan peninsula of Mexico using conventional culture methods (11 patients), and an immunocolorimetric detection of pathogenic Streptococcus pyogenes (15 patients). Prevalence of bacteria isolated by culture methods was 90.9% (10/11). We cultured, from chiclero's ulcers (60%), pathogenic bacterial such as Staphylococcus aureus (20%), S. pyogenes (1.6%), Pseudomonas aeruginosa (1.6%), Morganella morganii (1.6%), and opportunist pathogenic bacteria such as Klebsiella spp. (20.0%), Enterobacter spp. (20%), and Enterococcus spp. (20%). We also cultured coagulase-negative staphylococci in 40% (4/10) of the remaining patients. Micrococcus spp. and coagulase-negative staphylococci constituted the bacterial genuses more frequently isolated in the normal skin of patients with chiclero's ulcer and healthy individuals used as controls. We also undertook another study to find out the presence of S. pyogenes by an immunocolorimetric assay. This study indicated that 60% (9/15) of the ulcerated lesions, but not normal controls, were contaminated with S. pyogenes. Importantly, individuals with purulent secretion and holding concomitant infections with S. pyogenes, S. aureus, P. aeruginosa, M. morganii, and E. durans took longer to heal Leishmania (L.) mexicana infections treated with antimonial drugs. Our results suggest the need to eliminate bacterial purulent infections, by antibiotic treatment, before starting antimonial administration to patients with chiclero's ulcer.

Antimicrobial activity and chemical investigation of Brazilian Drosera

The antimicrobial activity of three different extracts (hexanic, ethyl acetate, methanol) obtained from Brazilian Drosera species (D. communis, D. montana var. montana, D. brevifolia, D. villosa var. graomogolensis, D. villosa var. villosa, Drosera sp. 1, and Drosera sp. 2 ) were tested against Staphylococcus aureus (ATCC 25923), Enterococcus faecium (ATCC23212), Pseudomonas aeruginosa (ATCC27853), Escherichia coli (ATCC11229), Salmonella choleraesuis (ATCC10708), Klebsiella pneumoniae (ATCC13883), and Candida albicans (a human isolate). Better antimicrobial activity was observed with D. communis and D. montana var. montana ethyl acetate extracts. Phytochemical analyses from D. communis, D. montana var. montana and D. brevifolia yielded 5-hydroxy-2-methyl-1,4-naphthoquinone (plumbagin); long chain aliphatic hydrocarbons were isolated from D. communis and from D. villosa var. villosa, a mixture of long chain aliphatic alcohols and carboxylic acids, was isolated from D. communis and 3b-O-acetylaleuritolic acid from D. villosa var. villosa.