Bleeding reduction after topical application of tranexamic acid together with Betadine solution in total knee arthroplasty. A randomised controlled study.

INTRODUCTION: Topical application of tranexamic acid to the knee joint before closure in total knee arthroplasty reduces postoperative bleeding without increase in complication. However, it is unknown the effectiveness of topic TXA performed with other topical medications, like povidone-iodine solution. MATERIALS AND METHODS: One hundred and twenty-five patients were randomized to receive 100mL of povidone-iodine solution (control: group A) or 1.5 (group B) and 3.0 g (group C) of topical TXA in povidone-iodine solution applied into the knee before closure in total knee arthroplasty. RESULTS: The patients in the TXA groups had higher mean postoperative hemoglobin levels (P=0.01 and P=0.03 in groups B and C, respectively) and a reduced postoperative blood loss in the TXA groups (P=0.07 and P=0.09 in groups B and C, respectively). No significant complications were observed. DISCUSSION: In this study, topical application of tranexamic acid after total knee arthroplasty together with povidone-iodine solution results in higher postoperative hemoglobin levels and lower blood loss compared with those in the control group without other complications. LEVEL OF EVIDENCE: I - I: high-powered prospective randomized trial.

Is the combination of pyrazinamide plus rifampicin safe for treating latent tuberculosis infection in persons not infected by the human immunodeficiency virus?

SETTING: Nine public health care centres in four Spanish cities. OBJECTIVE: To evaluate the efficacy and safety of 2 months of rifampicin (R) plus pyrazinamide (Z) therapy (2RZ) compared with a 6-month course of isoniazid therapy (6H) for treating latent tuberculosis infection (LTBI). DESIGN: Multicentered, randomised, comparative and prospective trial conducted in HIV-seronegative contacts of infectious pulmonary TB cases. RESULTS: Of 352 individuals, 199 received 6H and 153 2RZ; 73% of contacts receiving 6H and 71% receiving 2RZ completed treatment (P = 0.73). Treatment interruption due to hepatotoxicity (ALT/AST > 5 times upper limit of normal) was observed in 10% of contacts in the 2RZ group and in 2.5% of the 6H group (P = 0.007). This higher than expected rate of hepatotoxicity in the 2RZ arm led to premature termination of the study. Severe or fatal liver injury was not detected. Liver function tests normalised after discontinuation of treatment. We conclude that the use of RZ should only be considered when other regimens are unsuitable and intensive monitoring of liver function is feasible.

Effect of Brazilian green propolis on the production of reactive oxygen species by stimulated neutrophils.

The activity of a crude ethanol extract of green propolis and its fractions obtained by partition with hexane, chloroform and n-butanol was assessed on luminol- and lucigenin- enhanced chemiluminescence (CL) produced by rabbit neutrophils (PMNs) stimulated with particles of serum-opsonized zymosan (OZ). The total production of reactive oxygen species (ROS) by PMNs was measured by the luminol-enhanced CL (LumCL) assay and the production of the superoxide anion (O2*-) by the lucigenin-enhanced CL (LucCL) assay. All evaluated propolis samples had inhibitory effect on the LumCL and LucCL, which was concentration dependent. The n-butanol and chloroform fractions displayed the highest inhibitory effect on the LumCL produced by PMNs stimulated with OZ, in comparison with both the ethanol extract and the hexane fraction. Besides, the hexane fraction was the one which presented the highest effect for the LucCL assay. Some isolated compounds from both n-butanol and chloroform fractions were also assessed, including kaempferide, isosakuranetin, aromadendrine-4'-methyl-ether and 3-prenyl-p-coumaric acid. Kaempferide presented the highest inhibitory effect on the LumCL in comparison with the other compounds. Moreover, under the conditions assessed, the studied green propolis samples and isolated compounds were not toxic to the rabbit PMNs.

Melamine deaminase and atrazine chlorohydrolase: 98 percent identical but functionally different.

The gene encoding melamine deaminase (TriA) from Pseudomonas sp. strain NRRL B-12227 was identified, cloned into Escherichia coli, sequenced, and expressed for in vitro study of enzyme activity. Melamine deaminase displaced two of the three amino groups from melamine, producing ammeline and ammelide as sequential products. The first deamination reaction occurred more than 10 times faster than the second. Ammelide did not inhibit the first or second deamination reaction, suggesting that the lower rate of ammeline hydrolysis was due to differential substrate turnover rather than product inhibition. Remarkably, melamine deaminase is 98% identical to the enzyme atrazine chlorohydrolase (AtzA) from Pseudomonas sp. strain ADP. Each enzyme consists of 475 amino acids and differs by only 9 amino acids. AtzA was shown to exclusively catalyze dehalogenation of halo-substituted triazine ring compounds and had no activity with melamine and ammeline. Similarly, melamine deaminase had no detectable activity with the halo-triazine substrates. Melamine deaminase was active in deamination of a substrate that was structurally identical to atrazine, except for the substitution of an amino group for the chlorine atom. Moreover, melamine deaminase and AtzA are found in bacteria that grow on melamine and atrazine compounds, respectively. These data strongly suggest that the 9 amino acid differences between melamine deaminase and AtzA represent a short evolutionary pathway connecting enzymes catalyzing physiologically relevant deamination and dehalogenation reactions, respectively.

Peritoneal effusion in a dog secondary to visceral mast cell tumor. A case report.

BACKGROUND: Mast cell tumor, one of the most common skin tumors in dogs, may also be found in visceral sites (mainly spleen and liver). When a visceral mast cell tumor is present, neoplastic mast cells may be found in any effusion secondary to the tumor. Therefore, the diagnosis may be made by cytologic analysis of the effusion. CASE: An 8-year-old, spayed, female Siberian husky presented with a peritoneal effusion secondary to a visceral mast cell tumor. Seven months earlier, the dog had presented with a cutaneous nodule diagnosed as a well-differentiated mast cell tumor. The peritoneal fluid was classified as a transudate. Numerous neoplastic mast cells were found in the effusion. Although the mast cell tumor presented with characteristics of the well-differentiated tumor, its biologic behavior was that of a malignant tumor. CONCLUSION: Care should be taken to evaluate the prognosis of mast cell tumors in dogs since their biologic behavior is extremely variable.

Characterization of the ecdysteroid UDP-glucosyltransferase (egt) gene of Anticarsia gemmatalis nucleopolyhedrovirus.

The Anticarsia gemmatalis nucelopolyhedrovirus (AgMNPV) egt gene was cloned, sequenced and its expression characterized by RT-PCR and western blot analysis. Sequence analysis of the gene indicated the presence of an open reading frame (ORF) of 1482 nucleotides, which codes for a polypeptide of 494 amino acids. ATATA box and a conserved regulatory sequence (CATT) found in other baculovirus early genes were present in the promoter region of the egt gene. A poly-A consensus sequence was present in the 3' untranslated region (3'-UTR) of the gene. Homology comparisons showed that the EGT protein of AgMNPV is most closely related (95.9% amino acid sequence identity) to the EGT from the Choristoneura fumiferana DEF nucleopolyhedrovirus (CfDEF). Transcriptional analysis of the AgMNPV egt gene showed that egt-specific transcripts can be detected both early and late in infection. The EGT protein was detected, by western blot analysis, in the intra- (from 12 to 48 h post-infection) and extra-cellular (from 12 to 96 h post-infection) fractions of infected insect cells. The AgMNPV Bgl II-F fragment, which has homology to the AcMNPV ie-1 gene, was cloned and used to cotransfect SF21 cells with the cloned AgMNPV egt gene. EGT activity was observed, suggesting that AgMNPV ie-1 can transactivate egt expression.

The Bradyrhizobium japonicum noeD gene: a negatively acting, genotype-specific nodulation gene for soybean.

Bradyrhizobium japonicum strain USDA 110 is restricted for nodulation by soybean genotype PI 417566. We previously reported the identification of a USDA 110 Tn5 mutant, strain D4.2-5, that had the ability to overcome nodulation restriction conditioned by PI 417566 (S. M. Lohrke, J. H. Orf, E. Martínez-Romero, and M. J. Sadowsky, Appl. Environ. Microbiol. 61:2378-2383, 1995). In this study, we report the cloning and characterization of the negatively acting DNA region mutated in strain D4.2-5 that is involved in the genotype-specific nodulation of soybean. The Tn5 integration site was localized to a 5.2-kb EcoRI fragment isolated from wild-type USDA 110 genomic DNA. Saturation Tn5 mutagenesis of this 5.2-kb region and DNA homogenitization studies indicated that a 0.9-kb DNA region was involved in the genotype-specific nodulation of PI 417566. A single open reading frame (ORF) of 474 nucleotides, encoding a predicted protein of 158 amino acids, was identified within this region by DNA sequencing. This ORF was named noeD. Computer comparisons with available data bases revealed no significant similarities between the noeD DNA or predicted amino acid sequence and any known genes or their products. However, comparisons done with the region upstream of noeD revealed a high degree of similarity (about 76% similarity and 62% identity) to the N-terminal regions of the Rhizobium leguminosarum bv. viciae and R. meliloti nodM genes, which have been postulated to encode a glucosamine synthase. Southern hybridization analysis indicated that noeD is not closely linked to the main or auxiliary nodulation gene clusters in B. japonicum and that both nodulation-restricted and -unrestricted B. japonicum serogroup 110 strains contain a noeD homolog. High-performance liquid chromatography and fast atom bombardment-mass spectrometry analyses of the lipo-chitin oligosaccharide (LCO) nodulation signals produced by an noeD mutant showed a higher level of acetylation than that found with wild-type USDA 110. These results suggest that specific LCO signal molecules may be one of the factors influencing nodulation specificity in this symbiotic system.

The atzABC genes encoding atrazine catabolism are located on a self-transmissible plasmid in Pseudomonas sp. strain ADP.

Pseudomonas sp. strain ADP initiates atrazine catabolism via three enzymatic steps, encoded by atzA, -B, and -C, which yield cyanuric acid, a nitrogen source for many bacteria. In-well lysis, Southern hybridization, and plasmid transfer studies indicated that the atzA, -B, and -C genes are localized on a 96-kb self-transmissible plasmid, pADP-1, in Pseudomonas sp. strain ADP. High-performance liquid chromatography analyses showed that cyanuric acid degradation was not encoded by pADP-1. pADP-1 was transferred to Escherichia coli strains at a frequency of 4.7 x 10(-2). This suggests a potential molecular mechanism for the dispersion of the atzABC genes to other soil bacteria.

The atrazine catabolism genes atzABC are widespread and highly conserved.

Pseudomonas strain ADP metabolizes the herbicide atrazine via three enzymatic steps, encoded by the genes atzABC, to yield cyanuric acid, a nitrogen source for many bacteria. Here, we show that five geographically distinct atrazine-degrading bacteria contain genes homologous to atzA, -B, and -C. The sequence identities of the atz genes from different atrazine-degrading bacteria were greater than 99% in all pairwise comparisons. This differs from bacterial genes involved in the catabolism of other chlorinated compounds, for which the average sequence identity in pairwise comparisons of the known members of a class ranged from 25 to 56%. Our results indicate that globally distributed atrazine-catabolic genes are highly conserved in diverse genera of bacteria.

Molecular basis of a bacterial consortium: interspecies catabolism of atrazine.

Pseudomonas sp. strain ADP contains the genes, atzA, -B, and -C, that encode three enzymes which metabolize atrazine to cyanuric acid. Atrazine-catabolizing pure cultures isolated from around the world contain genes homologous to atzA, -B, and -C. The present study was conducted to determine whether the same genes are present in an atrazine-catabolizing bacterial consortium and how the genes and metabolism are subdivided among member species. The consortium contained four or more bacterial species, but two members, Clavibacter michiganese ATZ1 and Pseudomonas sp. strain CN1, collectively mineralized atrazine. C. michiganese ATZ1 released chloride from atrazine, produced hydroxyatrazine, and contained a homolog to the atzA gene that encoded atrazine chlorohydrolase. C. michiganese ATZ1 stoichiometrically metabolized hydroxyatrazine to N-ethylammelide and contained genes homologous to atzB and atzC, suggesting that either a functional AtzB or -C catalyzed N-isopropylamine release from hydroxyatrazine. C. michiganese ATZ1 grew on isopropylamine as its sole carbon and nitrogen source, explaining the ability of the consortium to use atrazine as the sole carbon and nitrogen source. A second consortium member, Pseudomonas sp. strain CN1, metabolized the N-ethylammelide produced by C. michiganese ATZ1 to transiently form cyanuric acid, a reaction catalyzed by AtzC. A gene homologous to the atzC gene of Pseudomonas sp. strain ADP was present, as demonstrated by Southern hybridization and PCR. Pseudomonas sp. strain CN1, but not C. michiganese, metabolized cyanuric acid. The consortium metabolized atrazine faster than did C. michiganese individually. Additionally, the consortium metabolized a much broader set of triazine ring compounds than did previously described pure cultures in which the atzABC genes had been identified. These data begin to elucidate the genetic and metabolic bases of catabolism by multimember consortia.

The atzB gene of Pseudomonas sp. strain ADP encodes the second enzyme of a novel atrazine degradation pathway.

We previously reported the isolation of a 21.5-kb genomic DNA fragment from Pseudomonas sp. strain ADP, which contains the atzA gene, encoding the first metabolic step for the degradation of the herbicide atrazine (M. de Souza, L. P. Wackett, K. L. Boundy-Mills, R. T. Mandelbaum, and M. J. Sadowsky, Appl. Environ. Microbiol. 61:3373-3378, 1995). In this study, we show that this fragment also contained the second gene of the atrazine metabolic pathway, atzB. AtzB catalyzed the transformation of hydroxyatrazine to N-isopropylammelide. The product was identified by use of high-performance liquid chromatography, mass spectrometery, and nuclear magnetic resonance spectroscopy. Tn5 mutagenesis of pMD1 was used to determine that atzB was located 8 kb downstream of atzA. Hydroxyatrazine degradation activity was localized to a 4.0-kb ClaI fragment, which was subcloned into the vector pACYC184 to produce plasmid pATZB-2. The DNA sequence of this region was determined and found to contain two large overlapping divergent open reading frames, ORF1 and ORF2. ORF1 was identified as the coding region of atzB by demonstrating that (i) only ORF1 was transcribed in Pseudomonas sp. strain ADP, (ii) a Tn5 insertion in ORF2 did not disrupt function, and (iii) codon usage was consistent with ORF1 being translated. AtzB had 25% amino acid identity with TrzA, a protein that catalyzes a hydrolytic deamination of the s-triazine substrate melamine. The atzA and atzB genes catalyze the first two steps of the metabolic pathway in a bacterium that rapidly metabolizes atrazine to carbon dioxide, ammonia, and chloride.

Anti-inflammatory effect of glucose-mannose binding lectins isolated from Brazilian beans.

Selectins are essential for leukocyte recruitment in inflammation. Because of a lectin domain present in the selectin structure, we investigated the anti-inflammtory activity of six mannose-glucose binding lectins from brazilian beans: Dioclea guianensis-DguiL; D. grandiflora-DgL; Cratylia floribunda-CfL; D. violacea-D.vL; D. virgata-DvirL and Canavalia brasiliensis-ConBr. The lectins were injected intravenously (i.v.) into rats (0.1 and 1.0 mg/kg; 30 min before irritants) and its activities compared to E. coli endotoxin (LPS,30 mug/kg i.v.). Three lectins (DvL, CfL and DguiL), although less intense than LPS, inhibited the neutrophil migration induced by carrageenan (Cg, 300 mug) in a dose-dependent manner (0.1 and 1.0 mg/kg). DvL activity was reversed by 0.1 M alpha-D-methyl-mannoside (alpha-CH3), but not by 0.1 M alpha-D-galactose. The fMLP (44 ng)-induced neutrophil migration was also reduced by these lectins. Endotoxin contamination of lectin samples could be excluded since alpha-CH3 treatment reversed the DvL effect, but did not modify LPS inhibitory activity. Carrageenan (300 mug)-induced paw oedema was also reduced by LPS or lectin treatments. Conversely, none of the tested lectins inhibited dextran (Dex, 300 mug)-induced paw oedema, a classical leukocyte independent model, or zymosan (Zy, 1.0 mg)-induced peritonitis and paw oedema. LPS showed no effect upon Dex-induced paw oedema and barely reduced (25%) the oedematogenic effects of zymosan. As proposed for LPS, the lectin inhibitory activity was better observed on neutrophil-mediated inflammatory reactions. We speculate that the plant lectin antiinflammatory activity is probably due to a competitive blockage of a common leukocyte and/or endothelial selectin carbohydrate ligand.

Atrazine chlorohydrolase from Pseudomonas sp. strain ADP: gene sequence, enzyme purification, and protein characterization.

Pseudomonas sp. strain ADP metabolizes atrazine to carbon dioxide and ammonia via the intermediate hydroxyatrazine. The genetic potential to produce hydroxyatrazine was previously attributed to a 1.9-kb AvaI DNA fragment from strain ADP (M. L. de Souza, L. P. Wackett, K. L. Boundy-Mills, R. T. Mandelbaum, and M. J. Sadowsky, Appl. Environ. Microbiol. 61:3373-3378, 1995). In this study, sequence analysis of the 1.9-kb AvaI fragment indicated that a single open reading frame, atzA, encoded an activity transforming atrazine to hydroxyatrazine. The open reading frame for the chlorohydrolase was determined by sequencing to be 1,419 nucleotides and encodes a 473-amino-acid protein with a predicted subunit molecular weight of 52,421. The deduced amino acid sequence matched the first 10 amino acids determined by protein microsequencing. The protein AtzA was purified to homogeneity by ammonium sulfate precipitation and anion-exchange chromatography. The subunit and holoenzyme molecular weights were 60,000 and 245,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration chromatography, respectively. The purified enzyme in H2(18)O yielded [18O]hydroxyatrazine, indicating that AtzA is a chlorohydrolase and not an oxygenase. The most related protein sequence in GenBank was that of TrzA, 41% identity, from Rhodococcus corallinus NRRL B-15444R. TrzA catalyzes the deamination of melamine and the dechlorination of deethylatrazine and desisopropylatrazine but is not active with atrazine. AtzA catalyzes the dechlorination of atrazine, simazine, and desethylatrazine but is not active with melamine, terbutylazine, or desethyldesisopropylatrazine. Our results indicate that AtzA is a novel atrazine-dechlorinating enzyme with fairly restricted substrate specificity and contributes to the microbial hydrolysis of atrazine to hydroxyatrazine in soils and groundwater.

[Epidemiology of violence--an approach to the problem of violent death in contemporary Brazilian society--the case in Santa Catarina]. / Epidemiologia da violência--uma aproximação ao problema através da morte violenta na sociedade brasileira contemporânea--o caso de Santa Catarina.

This paper discusses the problem of violence and its expression upon mortality due to external causes. A few indicators are offered, which have been worked upon it to emphasise the importance of the theme. In a general way, the study demonstrates violent death has had its magnitude increased along the years, not only throughout Latin America but also in Brazil and in Santa Catarina.

Cloning, characterization, and expression of a gene region from Pseudomonas sp. strain ADP involved in the dechlorination of atrazine.

We previously identified a Pseudomonas sp. strain, ADP, which rapidly metabolized atrazine in liquid culture, agar plates, and soils (R. T. Mandelbaum, D. L. Allan, L. P. Wackett, Appl. Environ. Microbiol. 61:1451-1457, 1995). In this study, we report the cloning and partial characterization of a gene region from Pseudomonas sp. strain ADP that encodes atrazine degradation activity. A 22-kb EcoRI genomic DNA fragment, designated pMD1, was shown to encode atrazine dechlorination activity in Escherichia coli DH5 alpha. Atrazine degradation was demonstrated by a zone-clearing assay on agar medium containing crystalline atrazine and by chromatographic methods. A gene conferring the atrazine-clearing phenotype was subsequently subcloned as a 1.9-kb AvaI fragment in pACYC184, designated pMD4, and was expressed in E. coli. This result and random Tn5 mutagenesis established that the 1.9-kb AvaI fragment was essential for atrazine dechlorination. High-pressure liquid and thin-layer chromatographic analyses were used to rigorously establish that E. coli containing pMD4 degraded atrazine and accumulated hydroxyatrazine. Hydroxyatrazine was detected only transiently in E. coli containing pMD1. This is consistent with the idea that hydroxyatrazine is the first metabolite in atrazine degradation by Pseudomonas sp. strain ADP. A 0.6-kb ApaI-PstI fragment from pMD4, containing the putative atrazine chlorohydrolase gene, hybridized to DNA from atrazine-degrading bacteria isolated in Switzerland and Louisiana.(ABSTRACT TRUNCATED AT 250 WORDS)

HTLV-I and HTLV-II infections among HIV-1 seropositive patients in Sao Paulo, Brazil.

To estimate the presence of, and the risk factors for HTLV-I and HTLV-II infections among HIV-1 infected subjects in Sao Paulo, Brazil, a serosurvey was performed in 471 HIV-1 infected patients, including 216 intravenous drug addicts (IVDA), 229 homosexual/bisexual men, and 26 with other risk factors. Serum samples were screened for HTLV seroreactivity by ELISA; reactive samples were analyzed by Western Blot (WB), using whole HTLV-I lysate as antigen. To confirm and discriminate HTLV-I and HTLV-II infections, sera presenting any bands on WB were further analyzed by a WB containing recombinant HTLV-I and HTLV-II proteins (WB 2.3), and by enzyme immunoassays using synthetic peptides specific for envelope proteins (Synth-EIA). In 22 cases, cell samples were available for polymerase chain reaction (PCR) studies. On WB, 114 sera were reactive and, of these, 37 and 25 were concordantly positive on both WB 2.3 and Synth-EIA procedures for HTLV-I and HTLV-II specific antibodies, respectively; 37 specimens were negative on both assays, and 15 gave discordant or indeterminate results. PCR findings confirmed concordant results obtained in the discriminatory serological assays. The prevalence rates of HTLV-I and HTLV-II infections were 15.3% and 11.1% in IVDA, and 0.9% and 0.4% in homosexual/bisexual men, respectively. No case of HTLV-I/HTLV-II co-infection was found.

PIP: HTLV-I is associated with adult T-cell leukemia/lymphoma and a neurological disorder known as HTLV-I-associated myelopathy or tropical spastic paraparesis. HTLV-II was initially isolated from subjects with a T-cell variant of hairy cell leukemia, but its etiological role in that or other diseases is unclear. HTLV infections, like HIV, are transmitted sexually, via blood transfusion and contaminated needles, and from mother to infant. Many reports indicate that HTLVs are present in the same populations at risk for HIV-1, and the cofactorial role of HTLVs in AIDS progression has been suggested by in vitro studies and epidemiological data. The authors report findings from a serosurvey conducted among 216 HIV-seropositive male and female intravenous drug users (IVDU), 229 HIV-seropositive homosexual and bisexual men, and 7 HIV-seropositive men and women who had had multiple transfusions, and 19 HIV-seropositive heterosexual men with multiple partners to estimate the presence of and the risk factors for HTLV-I and HTLV-II infections among HIV-1 infected individuals in Sao Paulo, Brazil. 70.9% of the subjects were classified according to CDC criteria as having AIDS. ELISA, Western blot, and polymerase chain reaction methods were used. The prevalence rates of HTLV-I and HTLV-II infections were 15.3% and 11.1% in IVDUs, and 0.9% and 0.4% in homosexual and bisexual men, respectively. No case of HTLV-I/HTLV-II co-infection was observed.

Lack of effect of the benzodiazepine receptor antagonist Ro 15-1788 on ethanol-induced intoxication in mice.

The effects of 3, 6 and 48 mg/kg of the benzodiazepine receptor antagonist Ro 15-1788 on the ethanol-induced depressant action were evaluated in mice. These results support and extend previous findings in experimental animals and show that Ro 15-1788 in doses devoid of intrinsic effects, does not antagonize the motor impairment, hypnotic effect or lethality induced by ethanol.