Biological and phytochemical potential of Baccharis trimera (Less.) DC leaf extract on swine clinical isolates.

Phytochemical studies have shown extracts obtained from the aerial parts of Baccharis trimera (Less.) DC has antioxidant and antimicrobial activities and the potential to treat some diseases. This study investigated the phenolic compounds, antioxidant and antimicrobial activity, and phytochemical potential of B. trimera leaf extract obtained by decoction on ATCC standard bacterial strains and 23 swine clinical isolates. Water was used as an extraction solvent based on the principles of green chemistry and at a low cost. The decoction process resulted in an extract rich in phenolic compounds and a high capacity for scavenging DPPH and ABTS radicals. Phytochemical analysis of aqueous extracts was performed using HPLC-DAD, and high concentrations of chlorogenic, ferulic, caffeic, and cinnamic phenolic acids were found. Antimicrobial activity was observed against gram-negative bacteria. B. trimera aqueous extract may be a promising low-cost agent for prophylactic treatment against swine enteropathogens and contribute to reducing production costs.

Borophosphate glasses as active agents for antimicrobial hydrogels.

Herein we report the synthesis of transition-metal-free potassium borophosphate glasses and their application as bactericidal and bacteriostatic material. The antimicrobial activity was achieved through a simple change in the molar ratio of boron and phosphorus atoms, making borophosphate glass soluble in water. The glasses were analyzed by X-ray powder diffraction, Raman spectroscopy, laser-induced breakdown spectroscopy, and water absorption. The addition of a boron compound is required to obtain potassium-based phosphate glasses. Moreover, the change in the phosphorus and boron molar ratio (P/B), 2, 1 or 0.5 affects the glass solubilization in water, which increases with the phosphorus content. The glass materials were submitted to tests of biological activity against the bacteria Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa. These water-soluble borophosphate glasses were employed in the development of hydrogel formulations using Carbopol®. Phosphorous-rich samples at a concentration of 15 % (w/w) in hydrogel showed better antimicrobial activity against S. aureus and E. coli, when compared to other samples, including commercial alcohol hand sanitizer gel, with an average size of the inhibition halo of 24.02±1.43 and 19.24±1.63mm, respectively.

Fumonisin-containing diets decrease the metabolic activity of myenteric neurons in rats.

Fumonisins are naturally occurring mycotoxins that contaminate food for human and animal consumption. They have neurotoxic effects, but the mechanisms by which these toxins affect the nervous system are not fully known. In the present study, male Wistar rats were fed between 21 and 63 days of age with diets that contained fumonisins B1+B2 at 0, 1, and 4 mg/kg. The following variables were assessed: food consumption, growth, body weight gain, and blood parameters. Morphoquantitave analyses of the most metabolically active myenteric neurons were performed, detected by NADH-diaphorase activity. Nitrergic neurons were detected by NADPH-diaphorase activity. The fumonisin-containing diets did not significantly alter food consumption or the body or plasma parameters. These diets decreased the metabolic activity of jejunal myenteric neurons, reducing neuronal density of the most metabolic active neurons by 30.8% and the cell body area by 4.3%. The diets also decreased the cell body area of nitrergic neurons by 22.1%. The effects of fumonisin B1 on the respiratory metabolism of isolated mitochondria in the brain and liver were also assessed. A decrease in oxygen consumption up to a 29% in the brain and 38% in the liver was observed in mitochondrial isolates to which 50 µM fumonisin B1 was added. The decrease in respiratory activity that was triggered by exposure to fumonisins was related to the lower metabolic activity of myenteric neurons, which had a negative impact on neuroplasticity of the enteric nervous system.

Chemical composition, antifungal and antioxidant activity of essential oils from Baccharis dracunculifolia and Pogostemon cablin against Fusarium graminearum.

Fusarium graminearum Schw. is associated with diseases in seeds and seedlings, mainly in maize, and causes damage to crops and food production. We determined the chemical compositions of essential oils from Pogostemon cablin (Blanco) Benth and Baccharis dracunculifolia DC., and evaluated their antioxidant and antifungal activity against F. graminearum. The oils were applied to the fungus using disc diffusion technique at concentrations of 8.0, 4.0, 2.0 and 1.0 µL mL-1. Fungistatic effects of P. cablin oil were detectable at 8.0 µL mL-1, with 80.0% inhibition of fungal growth. This oil contains mono and sesquiterpenes that may be toxic to fungal cell structures. P. cablin oil also had antioxidant activity in free radical sequestration experiments, where as oil from B. dracunculifolia had limited effects. We conclude that essential oil from P. cablin has greater anti-phytopathogenic and antioxidant activity than that from B. dracunculifolia.

Chronic ingestion of deoxynivalenol-contaminated diet dose-dependently decreases the area of myenteric neurons and gliocytes of rats.

BACKGROUND: Deoxynivalenol (DON), a mycotoxin produced by Fusarium spp., is commonly found in cereals ingested by humans and animals. Its ingestion is correlated with hepatic, hematologic, renal, splenic, cardiac, gastrointestinal, and neural damages, according to dose, duration of exposure and species. In this work, the effects of the ingestion of DON-contaminated diet at concentrations considered tolerable for human and animal intake were assessed. METHODS: Male Wistar rats aging 21 days were allotted to five groups that were given, for 42 days, diets contaminated with different concentrations of DON (0, 0.2, 0.75, 1.75, and 2 mg kg-1 of chow). Food ingestion, bodyweight, oxidative status and morphometric analyses of gliocytes, and neurons of jejunal myenteric ganglia were recorded. KEY RESULTS: At these concentrations, there was no food rejection, decrease in bodyweight gain, changes in oxidative status, or loss of either neurons or gliocytes. However, DON decreased gliocyte area, general neuronal population, nitrergic, cholinergic and NADH-diaphorase positive subpopulations and, as a result, ganglion area. CONCLUSIONS & INFERENCES: It was concluded that, even in the absence of visible effect, DON exposure reduces cell body area of gliocytes and neurons of the myenteric plexus of the rat jejunum.

Sulfonated (1→6)-ß-d-Glucan (Lasiodiplodan): Preparation, Characterization and Bioactive Properties.

Sulfonated derivatives of lasiodiplodan (LAS-S) with different degrees of substitution (1.61, 1.42, 1.02 and 0.15) were obtained and characterized by Fourier-transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and thermal and solubility analyses. Antimicrobial, antioxidant and cytotoxic potential were also assessed. The sulfonation was confirmed by FTIR analysis with specific bands at 1250 cm-1 (S=O, strong asymmetrical stretching vibration) and at 810 cm-1 (C-O-S, symmetrical vibration associated with the C-O-SO3 group) in the sulfonated samples. SEM demonstrated that sulfonation promoted morphological changes on the surface of the biopolymer with heterogeneous fibrillary structures appearing along the surface following chemical modification. LAS-S showed high thermal stability, with mass loss due to oxidation at temperatures close to 460 °C. Sulfonation increased the solubility of LAS, and in addition, increased the antimicrobial activity, especially against Candida albicans (fungicidal) and Salmonella enterica Typhimurium (bacteriostatic). Native lasiodiplodan (LAS-N) showed higher OH˙ removal capacity, while LAS-S had higher ferric ion reducing antioxidant power (FRAP) potential. LAS-N and LAS-S did not demonstrate lethal cytotoxicity against wild and mutant strains of Saccharomyces cerevisiae. Samples with higher degree of substitution (1.42 and 1.61) showed lower potential to induce oxidative stress.

Coq7p relevant residues for protein activity and stability.

Coenzyme Q (Q) is an isoprenylated benzoquinone electron carrier required for electronic transport in the mitochondrial respiratory chain, shuttling electrons from complexes I and II to complex III. Q synthesis requires proteins termed Coq (Coq1-Coq11). Coq7p is part of the multimeric complex involved in Q synthesis catalyzing the hydroxylation of demethoxy-Q6 (DMQ6), the last monooxygenase step in Q synthesis with a catalytic center containing a carboxylate-bridged di-iron at the active site of the enzyme. Here we indicate a group of Coq7p residues that modulate protein activity: D53, R57, V111 and S114. R57, V111 and S114 are very conserved residues; V111 and S114 are present in separated communities of amino acid correlation analysis. The coq7 double mutant V111G/S114A and S114E show respiratory deficiency at non permissive temperature, DMQ6 accumulation and lower content of Q6. Therefore we conclude that phosphomimetic S114E inhibit Coq7p activity, and propose that S114 phosphorylation is required to move a non-structured loop of 25 amino acids between helix 2 and 3, and that affects the di-iron coordination in Coq7p catalytic center.

nde1 deletion improves mitochondrial DNA maintenance in Saccharomyces cerevisiae coenzyme Q mutants.

Saccharomyces cerevisiae has three distinct inner mitochondrial membrane NADH dehydrogenases mediating the transfer of electrons from NADH to CoQ (coenzyme Q): Nde1p, Nde2p and Ndi1p. The active site of Ndi1p faces the matrix side, whereas the enzymatic activities of Nde1p and Nde2p are restricted to the intermembrane space side, where they are responsible for cytosolic NADH oxidation. In the present study we genetically manipulated yeast strains in order to alter the redox state of CoQ and NADH dehydrogenases to evaluate the consequences on mtDNA (mitochondrial DNA) maintenance. Interestingly, nde1 deletion was protective for mtDNA in strains defective in CoQ function. Additionally, the absence of functional Nde1p promoted a decrease in the rate of H2O2 release in isolated mitochondria from different yeast strains. On the other hand, overexpression of the predominant NADH dehydrogenase NDE1 elevated the rate of mtDNA loss and was toxic to coq10 and coq4 mutants. Increased CoQ synthesis through COQ8 overexpression also demonstrated that there is a correlation between CoQ respiratory function and mtDNA loss: supraphysiological CoQ levels were protective against mtDNA loss in the presence of oxidative imbalance generated by Nde1p excess or exogenous H2O2. Altogether, our results indicate that impairment in the oxidation of cytosolic NADH by Nde1p is deleterious towards mitochondrial biogenesis due to an increase in reactive oxygen species release.

Over-expression of COQ10 in Saccharomyces cerevisiae inhibits mitochondrial respiration.

COQ10 deletion in Saccharomyces cerevisiae elicits a defect in mitochondrial respiration correctable by addition of coenzyme Q(2). Rescue of respiration by Q(2) is a characteristic of mutants blocked in coenzyme Q(6) synthesis. Unlike Q(6) deficient mutants, mitochondria of the coq10 null mutant have wild-type concentrations of Q(6). The physiological significance of earlier observations that purified Coq10p contains bound Q(6) was examined in the present study by testing the in vivo effect of over-expression of Coq10p on respiration. Mitochondria with elevated levels of Coq10p display reduced respiration in the bc1 span of the electron transport chain, which can be restored with exogenous Q(2). This suggests that in vivo binding of Q(6) by excess Coq10p reduces the pool of this redox carrier available for its normal function in providing electrons to the bc1 complex. This is confirmed by observing that extra Coq8p relieves the inhibitory effect of excess Coq10p. Coq8p is a putative kinase, and a high-copy suppressor of the coq10 null mutant. As shown here, when over-produced in coq mutants, Coq8p counteracts turnover of Coq3p and Coq4p subunits of the Q-biosynthetic complex. This can account for the observed rescue by COQ8 of the respiratory defect in strains over-producing Coq10p.

Saccharomyces cerevisiae coq10 null mutants are responsive to antimycin A.

Deletion of COQ10 in Saccharomyces cerevisiae elicits a respiratory defect characterized by the absence of cytochrome c reduction, which is correctable by the addition of exogenous diffusible coenzyme Q(2). Unlike other coq mutants with hampered coenzyme Q(6) (Q(6) ) synthesis, coq10 mutants have near wild-type concentrations of Q(6). In the present study, we used Q-cycle inhibitors of the coenzyme QH(2)-cytochrome c reductase complex to assess the electron transfer properties of coq10 cells. Our results show that coq10 mutants respond to antimycin A, indicating an active Q-cycle in these mutants, even though they are unable to transport electrons through cytochrome c and are not responsive to myxothiazol. EPR spectroscopic analysis also suggests that wild-type and coq10 mitochondria accumulate similar amounts of Q(6) semiquinone, despite a lower steady-state level of coenzyme QH(2)-cytochrome c reductase complex in the coq10 cells. Confirming the reduced respiratory chain state in coq10 cells, we found that the expression of the Aspergillus fumigatus alternative oxidase in these cells leads to a decrease in antimycin-dependent H(2)O(2) release and improves their respiratory growth.

Site-directed mutagenesis and structural modeling of Coq10p indicate the presence of a tunnel for coenzyme Q6 binding.

Coq10p is a protein required for coenzyme Q function, but its specific role is still unknown. It is a member of the START domain superfamily that contains a hydrophobic tunnel implicated in the binding of lipophilic molecules. We used site-directed mutagenesis, statistical coupling analysis and molecular modeling to probe structural determinants in the Coq10p putative tunnel. Four point mutations were generated (coq10-K50E, coq10-L96S, coq10-E105K and coq10-K162D) and their biochemical properties analysed, as well as structural consequences. Our results show that all mutations impaired Coq10p function and together with molecular modeling indicate an important role for the Coq10p putative tunnel.

ADCK3, an ancestral kinase, is mutated in a form of recessive ataxia associated with coenzyme Q10 deficiency.

Muscle coenzyme Q(10) (CoQ(10) or ubiquinone) deficiency has been identified in more than 20 patients with presumed autosomal-recessive ataxia. However, mutations in genes required for CoQ(10) biosynthetic pathway have been identified only in patients with infantile-onset multisystemic diseases or isolated nephropathy. Our SNP-based genome-wide scan in a large consanguineous family revealed a locus for autosomal-recessive ataxia at chromosome 1q41. The causative mutation is a homozygous splice-site mutation in the aarF-domain-containing kinase 3 gene (ADCK3). Five additional mutations in ADCK3 were found in three patients with sporadic ataxia, including one known to have CoQ(10) deficiency in muscle. All of the patients have childhood-onset cerebellar ataxia with slow progression, and three of six have mildly elevated lactate levels. ADCK3 is a mitochondrial protein homologous to the yeast COQ8 and the bacterial UbiB proteins, which are required for CoQ biosynthesis. Three out of four patients tested showed a low endogenous pool of CoQ(10) in their fibroblasts or lymphoblasts, and two out of three patients showed impaired ubiquinone synthesis, strongly suggesting that ADCK3 is also involved in CoQ(10) biosynthesis. The deleterious nature of the three identified missense changes was confirmed by the introduction of them at the corresponding positions of the yeast COQ8 gene. Finally, a phylogenetic analysis shows that ADCK3 belongs to the family of atypical kinases, which includes phosphoinositide and choline kinases, suggesting that ADCK3 plays an indirect regulatory role in ubiquinone biosynthesis possibly as part of a feedback loop that regulates ATP production.

Vegetative compatibility and molecular characterization of Fusarium graminearum isolates from the State of Paraná, Brazil

Fusarium graminearum isolates causing Fusarium head blight in wheat were collected in Brazil and analyzed by random amplified polymorphic DNA (RAPD) markers and vegetative compatibility grouping (VCG). Nitrate non-utilizing mutants (nit) from each isolate were paired to verify heterokaryon formation. Three VCGs were identified among F. graminearum isolates: VCG1 included F-2, F-3 and F-4 isolates; VCG2 included F-1, F-6 and F-9 isolates; VCG3 included F-5, F-7 and F-8 isolates. Based on PCR amplification with eight different primers, the isolates showed great genetic similarity among themselves. Dendrogram analysis demonstrated two RAPD groups: Group A, consisting of isolates F-2 and F-9, and Group B, composed of the remaining isolates. Results suggest the clonal origin of F. graminearum isolates.

Isolados de Fusarium graminearum, obtidos de espigas de trigo com sintomas de Giberela, foram analisados pela técnica do Polimorfismo de DNA Amplificado ao Acaso (RAPD) e pelos Grupos de Compatibilidade Vegetativa (GCV). Mutantes auxotróficos (nit) de cada isolado foram pareados em todas as combinações possíveis, para a formação de heterocários. Três GCVs foram identificados: GCV1, incluindo os isolados F-2, F-3 e F-4; GCV2, incluindo os isolados F-1, F-6 e F-9; e GCV3, formado pelos isolados F-5, F-7 e F-8. Dois grupos foram identificados com base nos marcadores de RAPD: o grupo A, formado pelos isolados F-2 e F-9, e o grupo B, composto pelos demais isolados, os quais apresentaram grande similaridade entre si. Os resultados sugerem a origem clonal dos isolados de F. graminearum analisados.

Inibição do desenvolvimento de fungos fitopatogênicos por detergente derivado de óleo da mamona (Ricinus communis) / The castor oil plant detergent (Ricinus communis) inhibits the asexual development of phytopathogenic fungi

No presente estudo avaliou-se o efeito fungitóxico do detergente derivado do óleo da mamona (Ricinus communis) sobre o desenvolvimento dos fitopatógenos: Pyricularia grisea, Fusarium graminearum e Colletotrichum lindemuthianum. Seis concentrações do detergente (12,5mL L-1 a 300mL L-1) foram, individualmente, incorporadas ao Meio Basal; a seguir, após inoculação fúngica, o crescimento radial dos micélios foi avaliado. A inibição total do desenvolvimento de C. lindemuthianum e P. grisea foi observada entre as concentrações de 50mL L-1 e 200mL L-1, respectivamente. Com base no crescimento miceliano das colônias de F. graminearum, a atividade antifúngica do detergente do óleo da mamona (DOM) determinou inibição variável entre 79,4 e 91 por cento para a raça F2 e entre 80,7 e 90,7 por cento para a raça F4. O detergente, nas concentrações de 100 a 300mL L-1, inibiu em 100 por cento a germinação de conídios de F. graminearum (raças F-4 e F-2). Os resultados demonstram nítida atividade antifúngica do detergente derivado do óleo da mamona sobre fitopatógenos.

In the present study the fungitoxic effect of the castor oil plant detergent (Ricinus communis) on the development of the phytopathogens Pyricularia grisea, Fusarium graminearum and Colletotrichum lindemuthianum was evaluated. Six concentrations of the detergent (12.5mL L-1 to 300mL L-1) had been, individually, incorporated to the Basal Medium. After fungi inoculations, the radial growth of mycelia were evaluated. Detergent at 50mL L-1 and 200mL L-1 inhibited completely the development of P. grisea and C. lindemuthianum, respectively. On the basis of the mycelial growth of F. graminearum, the fungitoxic activity of the castor oil plant detergent (DOM) determined inhibition in the range of 79.4 and 91 percent for the F2 race and 80.7 and 90.7 percent for the F4 race. Detergent at the concentrations of 100mL L-1 to 300mL L-1 inhibited in 100 percent the F. graminearum germination conidia (races F-4 and F-2). Results demonstrate the fungitoxic activity of the castor oil plant detergent on phytopathogenic fungi.

Genetic and molecular characterization of pathogenic isolates of Pyricularia grisea from wheat (Triticum aestivum Lam.) and triticale (x Triticosecale Wittmack) in the state of Paraná, Brazil.

Isolates of Pyricularia grisea from wheat (Triticum aestivum Lam.) and triticale (x Triticosecale Wittmack) spikes with blast symptoms were analyzed by classical (VCG) and molecular (RAPD) techniques. P. grisea mutants, unable to use sodium nitrate (nit) as nitrogen source, were obtained with potassium chlorate. For vegetative compatibility (VCG) tests, genetically complementary nit mutant pairs were inoculated in a medium with sodium nitrate as a single nitrogen source. P. grisea isolates were divided into two vegetative compatibility groups and two RAPD groups. Since vegetative compatible strains may mutually exchange genetic and cytoplasmatic material, the contribution of the parasexual cycle in the genetic variability of Brazilian P. grisea isolates is discussed.

Genetic and molecular characterization of pathogenic isolates of Pyricularia grisea from wheat (Triticum aestivum Lam) and triticale (x Triticosecale Wittmack) in the state of Paraná, Brazil

Se analizaron por técnicas clásica (VCG) y molecular (RAPD) aislamientos dePyricularia grisea de las espigas de trigo (Triticum aestivum Lam.) y triticale(x Triticosecale Wittmack) con los síntomas de piricularia o bruzone.Los mutantes de P. grisea, incapaces de utilizar nitratos como fuente denitrógeno (nit), fueron obtenidos con clorato de potasio. Para las pruebasde compatibilidad vegetativa (VCG) se inocularon los pares complementariosde mutantes nit en un medio con nitrato de sodio como única fuente denitrógeno. Los aislamientos de P. grisea fueron separados en dos gruposde compatibilidad vegetativa y dos de RAPD. Considerándose que losaislamientos compatibles de P. grisea pueden intercambiar materialesnucleares y citoplasmáticos entre sí, se discute la contribución del cicloparasexual en la variabilidad genética de los aislamientos brasileños deP. grisea(AU)

Isolates of Pyricularia grisea from wheat (Triticum aestivum Lam.) and triticale(x Triticosecale Wittmack) spikes with blast symptoms were analyzed byclassical (VCG) and molecular (RAPD) techniques. P. grisea mutants, unable touse sodium nitrate (nit) as nitrogen source, were obtained with potassiumchlorate. For vegetative compatibility (VCG) tests, genetically complementarynit mutant pairs were inoculated in a medium with sodium nitrate as a singlenitrogen source. P. grisea isolates were divided into two vegetativecompatibility groups and two RAPD groups. Since vegetative compatiblestrains may mutually exchange genetic and cytoplasmatic material, thecontribution of the parasexual cycle in the genetic variability of BrazilianP. grisea isolates is discussed(AU)

Cremophor EL stimulates mitotic recombination in uvsH//uvsH diploid strain of Aspergillus nidulans.

Cremophor EL is a solubilizer and emulsifier agent used in the pharmaceutical and foodstuff industries. The solvent is the principal constituent of paclitaxel's clinical formulation vehicle. Since mitotic recombination plays a crucial role in multistep carcinogenesis, the study of the recombinagenic potential of chemical compounds is of the utmost importance. In our research genotoxicity of cremophor EL has been studied by using an uvsH//uvsH diploid strain of Aspergillus nidulans. Since it spends a great part of its cell cycle in the G2period, this fungus is a special screening system for the study of mitotic recombination induced by chemical substances. Homozygotization Indexes (HI) for paba and bi markers from heterozygous B211//A837 diploid strain were determined for the evaluation of the recombinagenic effect of cremophor EL. It has been shown that cremophor EL induces increase in mitotic crossing-over events at nontoxic concentrations (0.05 and 0.075% v/v).

Cremophor EL stimulates mitotic recombination in uvsH//uvsH diploid strain of Aspergillus nidulans

Cremophor EL is a solubilizer and emulsifier agent used in the pharmaceutical and foodstuff industries. The solvent is the principal constituent of paclitaxel's clinical formulation vehicle. Since mitotic recombination plays a crucial role in multistep carcinogenesis, the study of the recombinagenic potential of chemical compounds is of the utmost importance. In our research genotoxicity of cremophor EL has been studied by using an uvsH//uvsH diploid strain of Aspergillus nidulans. Since it spends a great part of its cell cycle in the G2period, this fungus is a special screening system for the study of mitotic recombination induced by chemical substances. Homozygotization Indexes (HI) for paba and bi markers from heterozygous B211//A837 diploid strain were determined for the evaluation of the recombinagenic effect of cremophor EL. It has been shown that cremophor EL induces increase in mitotic crossing-over events at nontoxic concentrations (0.05 and 0.075 percent v/v).