Culturas de bacilos ácido-álcool resistentes isolados de hematófagos infectados em leprosos: evidências de se tratar do bacilo de Hansen

The A. summarises the history of his first culture of acidfast bacillus isolated directly from leprosy lesions (Sample José) and refers about two samples recovered from guinea pig and white rat inoculated with said culture. Then the A. completes his previous descriptions of four cultures of acidfast bacilli isolated by him from ticks (Amblyomma cajaennense and Boophilus microplus, two cultures from each species) infected experimentally in lepers. The A. having found specimens of two species of Triatomidae (Triatoma infestans and Panstrongylus megistus) naturally infected with HANSEN bacillus in huts habited by lepers in the State of Minas Gerais (Dec. 1942), started a series of experiments, using larvae and nymphs of T. infestans bred in laboratory at the Instituto Oswaldo Cruz, to infect in active cases of leprosy, in the city of Rio de Janeiro, could obtain two new samples of cultures of acid-fast bacilli (Ns. 6 and 7 of his set). In this papaer the A. studies the biological properties of said cultures, proving that Penicilin has not effect upon them, like other substances. The sulphuric and acetic acids were used to purify some of the cultures, with good results, the cultures becoming more rich and growing faster. Potassium hydroxide Sodium (10% solution) was also used with success to isolate and to purify the cultures, but it seems that it affects the bacilli in some way. In flud glycerinated media the majority of such cultures produce velum suitable for the preparation of antigens for skin tests and for therapeutical use. At last the A. says that he is becoming convinced that the HANSEN bacillus is in cause, especially after thee evidences of culturing the bacillus from one patient, in different opportunities.

Nylon wool column: a tool for obtaining monokine-rich eluates in the absence of serum

Diverse conditions for stimulating human mononuclear cells to release thymocyte costimulatory factors were tested for their contribution to the generation of supernatants high titers of these monokines. Activity titers increased with LPS concentration, reaching a plateau between 1 and 10 microng/ml. Indomethacin did not modify the monokine, but the assay for thymocyte costimulatory activity was substantially affected by inhibitory substances produced by the monocytes in the absence of indomethacin. The use of nylon wool columns to trap the cells was shown to be effective in raising cellular densities without decreasing activity titers. As result, the yield per cell could be maintained even in the absence of serum, an important step toward the goal of purifiying bioactive from crude broths.

Thyroid peroxidase activity is inhibited by amino acids

Normal in vitro thyroid peroxidase (TPO) iodide oxidation activity was completely inhibited by a hydrolyzed TPO preparation (0.15 mg/ml) or hydrolyzed bovine serum albumin (BSA, 0.2 mg/ml). A pancreatic hydrolysate of casein (trypticase peptone, 0.1 mg/ml) and some amino acids (cysteine, tryptophan and methionine, 50 µM each) also inhibited the TPO iodide oxidation reaction completely, whereas casamino acids (0.1 mg/ml), and tyrosine, phenylalanine and histidine (50 µM each) inhibited the TPO reaction by 54% or less. A pancreatic digest of gelatin (0.1 mg/ml) or any other amino acid (50 µM) tested did not significantly decrease TPO activity. The amino acids that impair iodide oxidation also inhibit the TPO albumin iodination activity. The inhibitory amino acids contain side chains with either sulfur atoms (cysteine and methionine) or aromatic rings (tyrosine, tryptophan, histidine and phenylalanine). Among the amino acids tested, only cysteine affected the TPO guaiacol oxidation reaction, producing a transient inhibition at 25 or 50 µM. The iodide oxidation inhibitory activity of cysteine, methionine and tryptophan was reversed by increasing iodide concentrations from 12 to 18 mM, while no such effect was observed when the cofactor (H2O2) concentration was increased. The inhibitory substances might interfere with the enzyme activity by competing with its normal substrates for their binding sites, binding to the free substrates or reducing their oxidized form.

Use of real-time PCR to evaluate two DNA extraction methods from food

The DNA extraction is a critical step in Genetically Modified Organisms analysis based on real-time PCR. In this study, the CTAB and DNeasy methods provided good quality and quantity of DNA from the texturized soy protein, infant formula, and soy milk samples. Concerning the Certified Reference Material consisting of 5% Roundup Ready® soybean, neither method yielded DNA of good quality. However, the dilution test applied in the CTAB extracts showed no interference of inhibitory substances. The PCR efficiencies of lectin target amplification were not statistically different, and the coefficients of correlation (R²) demonstrated high degree of correlation between the copy numbers and the threshold cycle (Ct) values. ANOVA showed suitable adjustment of the regression and absence of significant linear deviations. The efficiencies of the p35S amplification were not statistically different, and all R² values using DNeasy extracts were above 0.98 with no significant linear deviations. Two out of three R² values using CTAB extracts were lower than 0.98, corresponding to lower degree of correlation, and the lack-of-fit test showed significant linear deviation in one run. The comparative analysis of the Ct values for the p35S and lectin targets demonstrated no statistical significant differences between the analytical curves of each target.

Antagonistic activity expressed by Shigella sonnei: identification of a putative new bacteriocin

Bacteriocins are antibacterial, proteinaceous substances that mediate microbial dynamics. Bacteriocin production is a highly disseminated property among all major lineages of bacteria, including Shigella. In this paper, we addressed the purification and characterisation of a bacteriocin produced by a Shigella sonnei strain (SS9) isolated from a child with acute diarrhoea. The substance was purified through ammonium-sulphate precipitation and sequential steps of chromatography. The intracellular fraction obtained at 75% ammonium sulphate maintained activity following exposure to pH values from 1-11 and storage at -80ºC for more than two years and was inactivated by high temperatures and proteases. The molecular mass of the purified bacteriocin was determined by mass spectrometry to be 18.56 kDa. The N-terminal sequence of the bacteriocin did not match any other antibacterial proteins described. A putative new bacteriocin produced by S. sonnei has been detected. This bacteriocin may represent a newly described protein or a previously described protein with a newly detected function. Considering that SS9 expresses antagonism against other diarrhoeagenic bacteria, the bacteriocin may contribute to S. sonnei virulence and is potentially applicable to either preventing or controlling diarrhoeal disease.

TIMP-1 mediates the inhibitory effect of interleukin-6 on the proliferation of a hepatocarcinoma cell line in a STAT3-dependent manner

The tissue inhibitor of metalloproteinases (TIMP)-1 is a multifunctional protein which is not only an inhibitor of matrix metalloproteinases (MMPs) but also to have a possible "cytokine-like" action. Here, we first compared mRNA expression of TIMP-1 and MMP-9 in BEL-7402 (a hepatocellular carcinoma cell line), L-02 (a normal liver cell line) and QSG-7701 (a cell line derived from peripheral tissue of liver carcinoma) using real-time quantitative RT-PCR. By evaluating the variation of the MMP-9/TIMP-1 ratio as an index of reciprocal changes of the expression of the two genes, we observed that the MMP-9/TIMP-1 ratio was about 13- and 5-fold higher in BEL-7402 than in L-02 and QSG-7701, respectively. Significantly, overexpression of TIMP-1 decreased the MMP-9/TIMP-1 ratio in BEL-7402 and then inhibited the cell growth to 60% and reduced the migration to about 30%. Meanwhile, our data showed that interleukin-6 (IL-6) (100 ng/mL) could also inhibited the cell growth of BEL-7402. Further studies indicated that TIMP-1 mediated the inhibitory effect of IL-6 on BEL-7402 cell proliferation in a STAT3-dependent manner, which could further accelerate the expression of the cyclin-dependent kinase inhibitor p21. A dominant negative STAT3 mutant totally abolished IL-6-induced TIMP-1 expression and its biological functions. The present results demonstrate that TIMP-1 may be one of the mediators that regulate the inhibitory effect of IL-6 on BEL-7402 proliferation in which STAT3 signal transduction and p21 up-regulation also play important roles.

Chlorpheniramine facilitates inhibitory avoidance in teleosts submitted to telencephalic ablation

The present study investigated the involvement of H(1) histaminegic receptor on the acquisition of inhibitory avoidance in Carassius auratus submitted to telencephalic ablation. The fish were submitted to telencephalic ablation 5 days before the experiment. The inhibitory avoidance procedure included 1 day for habituation, 3 days for training composed of 3 trials each (1st day: T1, T2, T3; 2nd day: 2T1, 2T2, 2T3; 3rd day: 3T1, 3T2, 3T3) and 1 day for test. On training days, the fish were placed in a white compartment, after 30 s the door was opened. When the fish crossed to a black compartment, a weight was dropped (aversive stimuli). Immediately after the third trial, on training days, the fish received, intraperitoneally, one of the pharmacological treatments (saline (N = 20), 8 (N = 12) or 16 (N = 13) µg/g chlorpheniramine, CPA). On the test day, the time to cross to the black compartment was determined. The latency of the saline group increased significantly only on the 3rd trial of the 2nd training day (mean ± SEM, T1 (50.40 ± 11.69), 2T3 (226.05 ± 25.01); ANOVA: P = 0.0249, Dunn test: P < 0.05). The group that received 8 µg/g CPA showed increased latencies from the 2nd training day until the test day (T1 (53.08 ± 17.17), 2T2 (197.75 ± 35.02), test (220.08 ± 30.98); ANOVA: P = 0.0022, Dunn test: P < 0.05)). These results indicate that CPA had a facilitating effect on memory. We suggest that the fish submitted to telencephalic ablation were able to learn due to the local circuits of the mesencephalon and/or diencephalon and that CPA interferes in these circuits, probably due an anxiolytic-like effect.

In vitro inhibitory effects of imatinib mesylate on stromal cells and hematopoietic progenitors from bone marrow

Imatinib mesylate (IM) is used to treat chronic myeloid leukemia (CML) because it selectively inhibits tyrosine kinase, which is a hallmark of CML oncogenesis. Recent studies have shown that IM inhibits the growth of several non-malignant hematopoietic and fibroblast cells from bone marrow (BM). The aim of the present study was to evaluate the effects of IM on stromal and hematopoietic progenitor cells, specifically in the colony-forming units of granulocyte/macrophage (CFU-GM), using BM cultures from 108 1.5- to 2-month-old healthy Swiss mice. The results showed that low concentrations of IM (1.25 µM) reduced the growth of CFU-GM in clonogenic assays. In culture assays with stromal cells, fibroblast proliferation and α-SMA expression by immunocytochemistry analysis were also reduced in a concentration-dependent manner, with a survival rate of approximately 50% with a dose of 2.5 µM. Cell viability and morphology were analyzed using MTT and staining with acrydine orange/ethidium bromide. Most cells were found to be viable after treatment with 5 µM IM, although there was gradual growth inhibition of fibroblastic cells while the number of round cells (macrophage-like cells) increased. At higher concentrations (15 µM), the majority of cells were apoptotic and cell growth ceased completely. Oil red staining revealed the presence of adipocytes only in untreated cells (control). Cell cycle analysis of stromal cells by flow cytometry showed a blockade at the G0/G1 phases in groups treated with 5-15 µM. These results suggest that IM differentially inhibits the survival of different types of BM cells since toxic effects were achieved.

Induction of chagasic-like arrhythmias in the isolated beating hearts of healthy rats perfused with Trypanosoma cruzi-conditioned medium

Chagas' myocardiopathy, caused by the intracellular protozoan Trypanosoma cruzi, is characterized by microvascular alterations, heart failure and arrhythmias. Ischemia and arrythmogenesis have been attributed to proteins shed by the parasite, although this has not been fully demonstrated. The aim of the present investigation was to study the effect of substances shed by T. cruzi on ischemia/reperfusion-induced arrhythmias. We performed a triple ischemia-reperfusion (I/R) protocol whereby the isolated beating rat hearts were perfused with either Vero-control or Vero T. cruzi-infected conditioned medium during the different stages of ischemia and subsequently reperfused with Tyrode's solution. ECG and heart rate were recorded during the entire experiment. We observed that triple I/R-induced bradycardia was associated with the generation of auricular-ventricular blockade during ischemia and non-sustained nodal and ventricular tachycardia during reperfusion. Interestingly, perfusion with Vero-infected medium produced a delay in the reperfusion-induced recovery of heart rate, increased the frequency of tachycardic events and induced ventricular fibrillation. These results suggest that the presence of parasite-shed substances in conditioned media enhances the arrhythmogenic effects that occur during the I/R protocol.

Effects of sciatic nerve transection on ultrastructure, NADPH-diaphorase reaction and serotonin-, tyrosine hydroxylase-, c-Fos-, glucose transporter 1- and 3-like immunoreactivities in frog dorsal root ganglion

Frogs have been used as an alternative model to study pain mechanisms. Since we did not find any reports on the effects of sciatic nerve transection (SNT) on the ultrastructure and pattern of metabolic substances in frog dorsal root ganglion (DRG) cells, in the present study, 18 adult male frogs (Rana catesbeiana) were divided into three experimental groups: naive (frogs not subjected to surgical manipulation), sham (frogs in which all surgical procedures to expose the sciatic nerve were used except transection of the nerve), and SNT (frogs in which the sciatic nerve was exposed and transected). After 3 days, the bilateral DRG of the sciatic nerve was collected and used for transmission electron microscopy. Immunohistochemistry was used to detect reactivity for glucose transporter (Glut) types 1 and 3, tyrosine hydroxylase, serotonin and c-Fos, as well as nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-diaphorase). SNT induced more mitochondria with vacuolation in neurons, satellite glial cells (SGCs) with more cytoplasmic extensions emerging from cell bodies, as well as more ribosomes, rough endoplasmic reticulum, intermediate filaments and mitochondria. c-Fos immunoreactivity was found in neuronal nuclei. More neurons and SGCs surrounded by tyrosine hydroxylase-like immunoreactivity were found. No change occurred in serotonin- and Glut1- and Glut3-like immunoreactivity. NADPH-diaphorase occurred in more neurons and SGCs. No sign of SGC proliferation was observed. Since the changes of frog DRG in response to nerve injury are similar to those of mammals, frogs should be a valid experimental model for the study of the effects of SNT, a condition that still has many unanswered questions.

Inhibitory effect of the essential oil from Eugenia caryophyllata Thumb leaves on coalho cheese contaminating microorganisms

Coalho cheese (a firm but very lightweight cheese produced in Brazil) is widely produced and consumed in the Brazilian Northeast and its production has been mainly related to small farmers. This food has been frequently characterized as having high microbial load posing a risk for the health of consumers. This study aimed to indentify the chemical compounds of the essential oil from Eugenia caryophyllata leaves; to evaluate the inhibitory effect of the oil against coalho cheese contaminating microorganisms; and to assess its efficacy in inhibiting the autochthonous microflora of the cheese during refrigerated storage. Eugenol (74%) was found to be the most prevalent compound in the essential oil. Minimum Inhibitory Concentration (MIC) and Minimum Cidal Concentration (MCC) in laboratorial broth were in the range of 2.5-5 and 5-20 µg.mL-1, respectively. Vaccum packed coalho cheese added with 5, 10, and 20 µg.g-1 of oil showed a lower growth rate (like-static effect) against mesophilic bacteria during the time intervals evaluated. On the other hand, 2.5-10 µg.g-1 of oil provided a prominent decrease toward fungi count in cheese samples during storage. These results reveal the interesting antimicrobial property of the essential oil from E. caryophyllata leaves against a range of coalho cheese-related microorganisms in laboratorial media and in food matrix.

What does the field of International Relations look like in South America?

Abstract This article provides a comprehensive picture of IR in South America by applying content analysis to 7,857 articles published in 35 journals from six South American countries from 2006 to 2014 in order to discover what the predominant theories, methods and research areas in this field are, how scholars tend to combine them in their research designs, and what the profiles of regional journals are, regarding their epistemological, methodological and subject preferences. The findings reveal a predominantly Positivist and largely Qualitative discipline, resembling North American and European IR.

Comparison of different methods for the determination of total organic carbon and humic substances in Brazilian soils

ABSTRACTAiming to compare three different methods for the determination of organic carbon (OC) in the soil and fractions of humic substances, seventeen Brazilian soil samples of different classes and textures were evaluated. Amounts of OC in the soil samples and the humic fractions were measured by the dichromate-oxidation method, with and without external heating in a digestion block at 130 °C for 30 min; by the loss-on-ignition method at 450 °C during 5 h and at 600 °C during 6 h; and by the dry combustion method. Dry combustion was used as reference in order to measure the efficiency of the other methods. Soil OC measured by the dichromate-oxidation method with external heating had the highest efficiency and the best results comparing to the reference method. When external heating was not used, the mean recovery efficiency dropped to 71%. The amount of OC was overestimated by the loss-on-ignition methods. Regression equations obtained between total OC contents of the reference method and those of the other methods showed relatively good adjustment, but all intercepts were different from zero (p < 0.01), which suggests that more accuracy can be obtained using not one single correction factor, but considering also the intercept. The Walkley-Black method underestimated the OC contents of the humic fractions, which was associated with the partial oxidation of the humin fraction. Better results were obtained when external heating was used. For the organic matter fractions, the OC in the humic and fulvic acid fractions can be determined without external heating if the reference method is not available, but the humin fraction requires the external heating.

Detection of Astrovirus-like in diarrhoeic stool and its coexistence with Rotavirus

A clinical case is described in this paper in that a 5-month old baby girl severely malnourished and dehydrated presented a prolonged acute diarrhoea. No enteropathogenic bacteria or parasites were demonstrated. Virological study by electron microscopy (EM) showed that the patient shed both astrovirus-like and rotavirus in the watery stool as long as 12 days after the onset. Immune electron microscopy (IEM) performed with the patient serum revealed clumps of both viruses. It is suggest that this may be a case of mixed infection due to astrovirus-like and rotavirus.