RESUMEN
Fusarium fujikuroi and Fusarium proliferatum are two phylogenetically closely related species of the Gibberella fujikuroi species complex (GFC). In some cases, strains of these species can cross and produce a few ascospores. In this study, we analyzed 26 single ascospore isolates of an interspecific cross between F. fujikuroi C1995 and F. proliferatum D4854 for their ability to produce four secondary metabolites: gibberellins (GAs), the mycotoxins fusarin C and fumonisin B(1), and a family of red polyketides, the fusarubins. Both parental strains contain the biosynthetic genes for all four metabolites, but differ in their ability to produce these metabolites under certain conditions. F. fujikuroi C1995 produces GAs and fusarins, while F. proliferatum D4854 produces fumonisins and fusarubins. The segregation amongst the progeny of these traits is not the expected 1:1 Mendelian ratio. Only eight, six, three and three progeny, respectively, produce GAs, fusarins, fumonisin B(1) and fusarubins in amounts similar to those synthesized by the producing parental strain. Beside the eight highly GA(3)-producing progeny, some of the progeny produce small amounts of GAs, predominantly GA(1), although these strains contain the GA gene cluster of the non-GA-producing F. proliferatum parental strain. Some progeny had recombinant secondary metabolite profiles under the conditions examined indicating that interspecific crosses can yield secondary metabolite production profiles that are atypical of the parent species.
Asunto(s)
Cruzamientos Genéticos , Fusarium/genética , Genes Fúngicos , Redes y Vías Metabólicas/genética , Recombinación Genética , Giberelinas/genética , Giberelinas/metabolismo , Micotoxinas/genética , Micotoxinas/metabolismo , Fenotipo , Policétidos/metabolismoRESUMEN
The authors studied the landscape components that favour the occurrence of anthrax in the Flooding Pampa grasslands (Buenos Aires province, Argentina). They made spatial locations of anthrax outbreaks diagnosed by registered veterinary laboratories in the study area's zone of influence. As variables for study, they differentiated areas that are flooded for 20% of the time or more from primary and secondary runoff channels. They also identified areas with low-productivity pasture. Logistic regression analysis of farm populations revealed that landscape components favouring the occurrence of anthrax outbreaks are shared runoff channels (odds ratio (OR) = 2.3; confidence interval (CI) = 1.2; 4.7) and > or = 40% low-productivity pasture (OR = 5.4; CI = 3.5; 8.3). Contrary to initial assumptions, susceptibility to flooding was not a significant variable (OR = 1.1; CI = 0.5; 2.1). The authors concluded that the first step in decision-making and ensuring more efficient implementation of future anthrax control and eradication plans was to identify risk variables.
Asunto(s)
Carbunco/veterinaria , Brotes de Enfermedades/veterinaria , Animales , Carbunco/epidemiología , Carbunco/etiología , Argentina/epidemiología , Intervalos de Confianza , Inundaciones , Modelos Logísticos , Oportunidad Relativa , Factores de RiesgoRESUMEN
The effect of grape seed extract (GS; 0.01% and 0.02%), oleoresin rosemary (OR; 0.02%) and water-soluble oregano extract (WS; 0.02%) on oxidative and color stability of cooked beef and pork patties stored at 4 degrees C for 8 d was determined. Fresh beef or pork lean and trim were ground, mixed (30% fat), and divided into 5 portions. Antioxidants mixed with salt (2%) were added. Patties were formed, cooked to an internal temperature of 71 degrees C, overwrapped in PVC, and stored at 4 degrees C. Lipid oxidation, assessed using thiobarbituric acid reactive substances (TBARS) and sensory evaluation, instrumental and visual color, and pH were determined after 0, 2, 4, 6, and 8 d. Based on TBARS values and off-odors associated with lipid oxidation such as rancidity, wet cardboard (for beef patties), and grassy (for beef and pork patties), grape seed extract resulted in the best antioxidant activity in both meat species. It did not change instrumental color measures of redness, yellowness, or color intensity, and appeared to reduce visual green discoloration in beef patties. The higher GS concentration (0.02%) exhibited more antioxidant activity than the lower concentration (0.01%). Therefore, grape seed extract at 0.02% has the potential to reduce oxidative rancidity and improve shelf life of refrigerated cooked beef and pork patties.
Asunto(s)
Antioxidantes/farmacología , Culinaria/métodos , Carne , Refrigeración/métodos , Animales , Bovinos , Manipulación de Alimentos/métodos , Concentración de Iones de Hidrógeno , Metabolismo de los Lípidos , Odorantes , Origanum , Oxidación-Reducción/efectos de los fármacos , Pigmentos Biológicos , Extractos Vegetales/farmacología , Control de Calidad , Rosmarinus , Cloruro de Sodio/administración & dosificación , Porcinos , Gusto , Sustancias Reactivas al Ácido Tiobarbitúrico/metabolismo , Factores de Tiempo , VitisRESUMEN
The objective of this study was to evaluate the efficacy of shelf-life enhancers (sodium lactate [SL] or sodium lactate+sodium diacetate [SLDA]) on limiting the growth of Escherichia coli K12 in needle-injecting surface-contaminated beef strip steaks with recycled enhancement solutions. Strip loins were injected to 10% over initial weight. SLDA was most effective in controlling bacterial growth followed by SL alone. The salt/phosphate combination in the enhancement solution was ineffective. SL decreased a(∗) (less red) and b(∗) values (less yellow) of the steaks compared to the salt/phosphate control. SLDA decreased L(∗) values compared to SL alone. SL and SLDA were equally effective in reducing purge loss from the steaks. Steaks enhanced with a "fresh" enhancement solution had higher a(∗) values (more red) and higher E. coli contamination levels than steaks enhanced with recycled solutions. Recycling did not affect L(∗) value, b(∗) value, or purge loss.
RESUMEN
The objective of this study was to evaluate the efficacy of shelf-life enhancers (sodium lactate [SL] or sodium lactate + sodium diacetate [SLDA]) on limiting the growth of Escherichia coli K12 in needle-injecting surface-contaminated beef strip steaks with recycled enhancement solutions. Strip loins were injected to 10% over initial weight. SLDA was most effective in controlling bacterial growth followed by SL alone. The salt/phosphate combination in the enhancement solution was ineffective. SL decreased a(∗) (less red) and b(∗) values (less yellow) of the steaks compared to the salt/phosphate control. SLDA decreased L(∗) values compared to SL alone. SL and SLDA were equally effective in reducing purge loss from the steaks. Steaks enhanced with a "fresh" enhancement solution had higher a(∗) values (more red) and higher E. coli contamination levels than steaks enhanced with recycled solutions. Recycling did not affect L(∗) value, b(∗) value, or purge loss.
RESUMEN
Nine biological species, or mating populations (MPs), denoted by letters A to I, and at least 29 anamorphic Fusarium species have been identified within the Gibberella fujikuroi species complex. Members of this species complex are the only species of the genus Fusarium that contain the gibberellin (GA) biosynthetic gene cluster or at least parts of it. However, the ability of fusaria to produce GAs is so far restricted to Fusarium fujikuroi, although at least six other MPs contain all the genes of the GA biosynthetic gene cluster. Members of Fusarium proliferatum, the closest related species, have lost the ability to produce GAs as a result of the accumulation of several mutations in the coding and 5' noncoding regions of genes P450-4 and P450-1, both encoding cytochrome P450 monooxygenases, resulting in metabolic blocks at the early stages of GA biosynthesis. In this study, we have determined additional enzymatic blocks at the first specific steps in the GA biosynthesis pathway of F. proliferatum: the synthesis of geranylgeranyl diphosphate and the synthesis of ent-kaurene. Complementation of these enzymatic blocks by transferring the corresponding genes from GA-producing F. fujikuroi to F. proliferatum resulted in the restoration of GA production. We discuss the reasons for Fusarium species outside the G. fujikuroi species complex having no GA biosynthetic genes, whereas species distantly related to Fusarium, e.g., Sphaceloma spp. and Phaeosphaeria spp., produce GAs.
Asunto(s)
Diterpenos de Tipo Kaurano/biosíntesis , Fusarium/enzimología , Fusarium/genética , Prueba de Complementación Genética , Giberelinas/biosíntesis , Fosfatos de Poliisoprenilo/biosíntesis , Transferasas Alquil y Aril/química , Transferasas Alquil y Aril/genética , Transferasas Alquil y Aril/metabolismo , Secuencia de Aminoácidos , Sistema Enzimático del Citocromo P-450/química , Sistema Enzimático del Citocromo P-450/genética , Sistema Enzimático del Citocromo P-450/metabolismo , Diterpenos , Farnesiltransferasa/química , Farnesiltransferasa/genética , Farnesiltransferasa/metabolismo , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Oxigenasas de Función Mixta/química , Oxigenasas de Función Mixta/genética , Oxigenasas de Función Mixta/metabolismo , Datos de Secuencia Molecular , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Análisis de Secuencia de ADNRESUMEN
Gibberella fujikuroi is a species complex with at least nine different biological species, termed mating populations (MPs) A to I (MP-A to MP-I), known to produce many different secondary metabolites. So far, gibberellin (GA) production is restricted to Fusarium fujikuroi (G. fujikuroi MP-C), although at least five other MPs contain all biosynthetic genes. Here, we analyze the GA gene cluster and GA pathway in the closest related species, Fusarium proliferatum (MP-D), and demonstrate that the GA genes share a high degree of sequence homology with the corresponding genes of MP-C. The GA production capacity was restored after integration of the entire GA gene cluster from MP-C, indicating the existence of an active regulation system in F. proliferatum. The results further indicate that one reason for the loss of GA production is the accumulation of several mutations in the coding and 5' noncoding regions of the ent-kaurene oxidase gene, P450-4.
Asunto(s)
Sistema Enzimático del Citocromo P-450/genética , Fusarium/enzimología , Fusarium/genética , Genes Fúngicos , Gibberella/enzimología , Gibberella/genética , Familia de Multigenes , Oxigenasas/genética , Secuencia de Bases , Sistema Enzimático del Citocromo P-450/metabolismo , ADN de Hongos/genética , Genes Reguladores , Genes Reporteros , Giberelinas/biosíntesis , Datos de Secuencia Molecular , Oxigenasas/metabolismo , Regiones Promotoras Genéticas , Homología de Secuencia de Ácido Nucleico , Especificidad de la EspecieRESUMEN
The objective of this study was to characterize the immune response of Babesia bigemina-infected cows during the second trimester of pregnancy. Twelve animals were divided into four groups (I, II, III, IV); groups I and II were pregnant cows, groups III and IV were non-pregnant cows. Groups I and III were infected with a virulent strain of Babesia bigemina, the doses utilized was 1 x 10(7) infected red blood cells IM. Groups II and IV were noninfected control groups. All the infected animals were severely affected; at days 5-7 post-inoculation (DPI) they showed clinical signs: fever (40-41.5 degrees C), packed cell volume reduction, and parasitemia, and specific treatment was required. The immune response was monitored daily from 0-11 DPI. As shown by flow cytometry analysis, in infected animals the distribution in peripheral blood of the T-cells subpopulations (CD4+, CD8+, gammadelta T-cells) was not affected when compared to the control groups. By ELISA, IFN-gamma production showed a trend to increase in plasma between 6-10 DPI; noninfected cows showed the lowest optical density values. By RT-PCR, a Th1 predominant response was observed, TNFalpha, INF-gamma and iNOs were detected. In contrast IL-4 and IL-10 were weak or undetected. The results of this trial will be discussed.
Asunto(s)
Babesia/inmunología , Babesia/patogenicidad , Babesiosis/inmunología , Babesiosis/veterinaria , Enfermedades de los Bovinos/inmunología , Citocinas/inmunología , Animales , Formación de Anticuerpos , Bovinos , Citocinas/análisis , Citocinas/biosíntesis , Ensayo de Inmunoadsorción Enzimática , Femenino , Inflamación , Interferón gamma/análisis , Embarazo , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Babesiosis is a tick-borne disease of cattle caused by Babesia bigemina and Babesia bovis and is transmitted by the tick vector Boophilus microplus. In this study, we investigate B. bigemina infection regarding the clinical infection, T cell distribution, and cytokine profile during the acute phase of an experimental infection in pregnant cows.
Asunto(s)
Babesia/inmunología , Babesia/patogenicidad , Babesiosis/inmunología , Babesiosis/veterinaria , Enfermedades de los Bovinos/inmunología , Enfermedades de los Bovinos/parasitología , Linfocitos T/inmunología , Animales , Bovinos , Citocinas/análisis , Femenino , Embarazo , Enfermedades por Picaduras de Garrapatas/inmunología , Enfermedades por Picaduras de Garrapatas/veterinariaAsunto(s)
Drenaje/métodos , Hematoma/cirugía , Enfermedades Orbitales/cirugía , Niño , Femenino , Humanos , AgujasRESUMEN
Recent studies have shown that the genes of the gibberellin (GA) biosynthesis pathway in the fungus Gibberella fujikuroi are organized in a cluster of at least seven genes. P450-1 is one of four cytochrome P450 monooxygenase genes in this cluster. Disruption of the P450-1 gene in the GA-producing wild-type strain IMI 58289 led to total loss of GA production. Analysis of the P450-1-disrupted mutants indicated that GA biosynthesis was blocked immediately after ent-kaurenoic acid. The function of the P450-1 gene product was investigated further by inserting the gene into mutants of G. fujikuroi that lack the entire GA gene cluster; the gene was highly expressed under GA production conditions in the absence of the other GA-biosynthesis genes. Cultures of transformants containing P450-1 converted ent-[(14)C]kaurenoic acid efficiently into [(14)C]GA(14), indicating that P450-1 catalyzes four sequential steps in the GA-biosynthetic pathway: 7beta-hydroxylation, contraction of ring B by oxidation at C-6, 3beta-hydroxylation, and oxidation at C-7. The GA precursors ent-7alpha-hydroxy[(14)C]kaurenoic acid, [(14)C]GA(12)-aldehyde, and [(14)C]GA(12) were also converted to [(14)C]GA(14). In addition, there is an indication that P450-1 may also be involved in the formation of the kaurenolides and fujenoic acids, which are by-products of GA biosynthesis in G. fujikuroi. Thus, P450-1 displays remarkable multifunctionality and may be responsible for the formation of 12 products.
Asunto(s)
Sistema Enzimático del Citocromo P-450/genética , Gibberella/genética , Giberelinas/biosíntesis , Secuencia de Bases , Northern Blotting , Southern Blotting , Cartilla de ADN , Gibberella/enzimología , Reacción en Cadena de la PolimerasaRESUMEN
A microsomal preparation from mycelia of the gibberellin (GA)-producing fungus Gibberella fujikuroi catalyzed the first two steps in the conversion of the biosynthetic intermediate GA12-aldehyde to gibberellic acid (GA3). [14C]GA12-Aldehyde was converted to radiolabelled GA14, the major product, together with smaller amounts of non-hydroxylated GA12. The microsomal activities required reduced pyridine nucleotides and molecular oxygen. However, GA12 and GA14 synthesis differed markedly in the preferred electron source. Formation of GA12 required NADH or NADPH, while GA14 synthesis from GA12-aldehyde occurred only with NADPH. Marked differences were also found in the activating effect of FAD. When NADPH was the reductant, the rate of GA14 synthesis was enhanced 3.5 times by 5 microM FAD while this flavin nucleotide did not alter the synthesis of GA12. In contrast, GA12 synthesis was activated 3.8 times by 50 microM FAD in the presence of NADH. Both activities were inhibited by carbon monoxide and cytochrome c. These properties suggest that the 3beta-hydroxylation of GA12-aldehyde and further oxidation of carbon 7 are catalyzed by cytochrome P-450 monooxygenases in Gibberella fujikuroi.
Asunto(s)
Ascomicetos/metabolismo , Giberelinas/biosíntesis , Oxigenasas de Función Mixta/metabolismo , Ascomicetos/enzimología , Monóxido de Carbono/metabolismo , Grupo Citocromo c/antagonistas & inhibidores , Inhibidores Enzimáticos/farmacología , Oxigenasas de Función Mixta/antagonistas & inhibidoresRESUMEN
Saccharomyces cerevisiae (ATP) and cytosolic rat liver (GTP) phospho enol pyruvate carboxykinases (EC 4.1.1.49/32) have been labeled with N-(1-pyrenyl)-iodoacetamide. Reagent incorporation was completely prevented by the presence of the respective nucleoside diphosphate plus MnCl2. Under appropriate conditions, 2 mol of reagent per mol of enzyme subunit were incorporated. The fluorescence spectra of the labeled proteins showed the pyrene excimer emission band. The pyrenyl-derivatized enzymes were digested with trypsin after carboxymethylation, and two labeled peptides were isolated for each carboxykinase upon reverse-phase high-performance liquid chromatography. Automated Edman degradation of the labeled peptides indicated that cysteines 364 and 457 (yeast enzyme), and cysteines 288 and 413 (rat enzyme) were labeled with the fluorescence SH-specific reagent. The relative reactivity of these residues was characterized. Labeling experiments utilizing the 5,5'-dithiobis(2-nitrobenzoate)-oxidized enzymes suggested that the reactive SH-groups occupy a vicinal position in the tertiary structure of the proteins, probably in the nucleotide-binding region.
Asunto(s)
Cisteína/análisis , Glucósidos/análisis , Hígado/enzimología , Fosfoenolpiruvato Carboxiquinasa (GTP)/análisis , Pirimidinonas/análisis , Saccharomyces cerevisiae/metabolismo , Adenosina Trifosfato , Secuencia de Aminoácidos , Animales , Citosol/enzimología , Ácido Ditionitrobenzoico , Colorantes Fluorescentes , Guanosina Trifosfato , Yodoacetamida/análogos & derivados , Fosfoenolpiruvato Carboxiquinasa (GTP)/aislamiento & purificación , RatasRESUMEN
Evaluamos la inmunogenicidad de la vacuna cubana recombinante contra HBV aplicando el esquema 12 meses, en niños de 1 a 10 años, encontrándose que con la segunda dosis el 98 por ciento tienen seroprotección ) valores > o igual 10 UI/I) y después de la tercera dosis (75 días) el 100 por ciento de los vacunados tienen niveles de anticuerpos >o igual 100 UI/I(AU)
Asunto(s)
Humanos , Preescolar , Niño , Vacunas Sintéticas/inmunología , Vacunas contra Hepatitis B/inmunología , Evaluación de Medicamentos , Hepatitis B/prevención & controlRESUMEN
Evaluamos la inmunogenicidad de la vacuna cubana recombinante contra HBV aplicando el esquema 012 meses, en niños de 1 a 10 años, encontrándose que con la segunda dosis el 98 por ciento tienen seroprotección ) valores > o igual 10 UI/I) y después de la tercera dosis (75 días) el 100 por ciento de los vacunados tienen niveles de anticuerpos >o igual 100 UI/I
Asunto(s)
Niño , Vacunas Sintéticas/inmunología , Vacunas contra Hepatitis Viral , Hepatitis B/inmunologíaRESUMEN
Los resultados obtenidos por el método inmunoenzimático cuantitativo de Abbott al primer año de completados los esquemas 012 y 016 meses, con la vacuna recombinante cubana contra la hepatitis viral B, muestran la inmunogenicidad en los vacunados con los dos esquemas, viéndose que el 100 por ciento tienen seroprotección y el 97 por ciento de estos tienen valores >o igual 100 UI/I para ambos esquemas. En cuanto a los valores de las medias geométricas, observamos que no se evidencian disminuciones significativas con el esquema 012, pero sí con el esquema 016, por lo cual sugerimos el uso del esquema 012, debido a su corta latencia, simetría y estabilidad de los valores de anticuerpos anti-HBs
Asunto(s)
Niño , Vacunas Sintéticas/uso terapéutico , Vacunas contra Hepatitis Viral , Hepatitis BRESUMEN
Two members of the ATP-dependent class of phospho enol pyruvate (PEP) carboxykinases (Saccharomyces cerevisiae and Escherichia coli PEP carboxykinase), and one member of the GTP-dependent class (the cytosolic rat liver enzyme) have been comparatively analyzed by taking advantage of their intrinsic fluorescence. The S. cerevisiae and the rat liver enzymes show intrinsic fluorescence with a maximum emission characteristic of moderately buried tryptophan residues, while the E. coli carboxykinase shows somewhat more average exposure for these fluorophores. The fluorescence of the three proteins was similarly quenched by the polar compound acrylamide, but differences were observed for the ionic quencher iodide. For the ATP-dependent enzymes, these last experiments indicate more exposure to the aqueous media of the tryptophan population of the E. coli than of the S. cerevisiae enzyme. The effect of nucleotides on the emission intensities and quenching efficiencies revealed substrate-induced conformational changes in the E. coli and cytosolic rat liver PEP carboxykinases. The addition of Mn2+ or of the adenosine nucleotides in the presence of Mg2+ induced an enhancement in the fluorescence of the E. coli enzyme. The addition of guanosine or inosine nucleotides to the rat liver enzyme quenched its fluorescence. From the ligand-induced fluorescence changes, dissociation constants of 40 +/- 6 microM, 10 +/- 0.8 microM, and 15 +/- 1 microM were obtained for Mn2+, MgATP and MgADP binding to the E. coli enzyme, respectively. For the cytosolic rat liver PEP carboxykinase, the respective values for GDP, IDP and ITP binding are 6 +/- 0.5 microM, 6.7 +/- 0.4 microM and 10.1 +/- 1.7 microM. A comparison of the dissociation constants obtained in this work with those reported for other PEP carboxykinases is presented.
Asunto(s)
Escherichia coli/enzimología , Hígado/enzimología , Fosfoenolpiruvato Carboxiquinasa (GTP)/química , Saccharomyces cerevisiae/enzimología , Triptófano/análisis , Acrilamida , Acrilamidas , Adenosina Difosfato/metabolismo , Adenosina Trifosfato/metabolismo , Adenosina Trifosfato/farmacología , Animales , Sitios de Unión , Citosol/enzimología , Fluorescencia , Guanosina Trifosfato/farmacología , Manganeso , Nucleótidos/farmacología , Fosfoenolpiruvato Carboxiquinasa (GTP)/metabolismo , Conformación Proteica , Ratas , Yoduro de SodioRESUMEN
Este estudio compara la inmunogenicidad (seroconversión, seroprotección e hiperrespuesta), producida por dos vacunas recombinantes contra la hepatitis B (Engerix-B de Bélgica y Cubana), en dos esquemas (012 y 016 meses), empleando los métodos de cuantificación para anti-HBsAg (Abbott y Organón), los cuales fueron también comparados. En el estudio participaron 257 voluntarios, divididos al azar en 4 grupos (dos vacunas, dos esquemas). Resultados: los dos métodos de Abbott y Organon, no presentan diferencias estadísticas significativas. La vacuna cubana muestra una mayor respuesta inmunogénica para dos dosis de vacuna y para el esquema 012. No hay diferencia entre los esquemas 012 y 016 y en el esquema 016 no se ven diferencias estadísticamente significativas con la vacuna Engerix-B. En esta última el esquema 016 muestra mejores resultados que el 012
Asunto(s)
Vacunas contra Hepatitis B/inmunología , Vacunas Sintéticas/inmunologíaRESUMEN
Reaction of rat liver phosphoenolpyruvate carboxykinase (GTP: oxaloacetate carboxy-lyase (transphosphorylating), EC 4.1.1.32) with the alkylating fluorescent probe N-(iodoacetylaminoethyl)-5-naphthylamine-1-sulfonic acid (1,5-I-AEDANS), results in complete loss of enzymatic activity. One mole of the fluorescent reagent is incorporated per mole of the inactivated enzyme. When the modification is carried out in the presence of GDPMn, the enzyme retains 97% of its activity with almost no incorporation of label. The specificity of the reaction is further supported by the detection of a unique fluorescent peptide from the trypsin-treated modified enzyme. Fluorescence emission of enzyme-bound AEDANS shows a broad band centered at 470 nm and presents a monoexponential decay with a lifetime of 19 ns. These data indicate that the probe-binding site is considerably less polar than water and similar in polarity to ethanol. Anisotropy determinations give evidence for restricted rotational freedom for AEDANS bound to the rat carboxykinase, while acrylamide quenching studies reveal limited accessibility to the probe site. The results are consistent with specific labeling of rat liver phosphoenolpyruvate carboxykinase at or near the GDP site. The characteristics of the nucleotide-binding sites of rat liver and yeast (ATP) phosphoenolpyruvate carboxykinase are compared.