RESUMEN
BACKGROUND: Theileria parva causes East Coast fever (ECF), one of the most economically important tick-borne diseases of cattle in sub-Saharan Africa. A live immunisation approach using the infection and treatment method (ITM) provides a strong long-term strain-restricted immunity. However, it typically induces a tick-transmissible carrier state in cattle and may lead to spread of antigenically distinct parasites. Thus, understanding the genetic composition of T. parva is needed prior to the use of the ITM vaccine in new areas. This study examined the sequence diversity and the evolutionary and biogeographical dynamics of T. parva within the African Great Lakes region to better understand the epidemiology of ECF and to assure vaccine safety. Genetic analyses were performed using sequences of two antigen-coding genes, Tp1 and Tp2, generated among 119 T. parva samples collected from cattle in four agro-ecological zones of DRC and Burundi. RESULTS: The results provided evidence of nucleotide and amino acid polymorphisms in both antigens, resulting in 11 and 10 distinct nucleotide alleles, that predicted 6 and 9 protein variants in Tp1 and Tp2, respectively. Theileria parva samples showed high variation within populations and a moderate biogeographical sub-structuring due to the widespread major genotypes. The diversity was greater in samples from lowlands and midlands areas compared to those from highlands and other African countries. The evolutionary dynamics modelling revealed a signal of selective evolution which was not preferentially detected within the epitope-coding regions, suggesting that the observed polymorphism could be more related to gene flow rather than recent host immune-based selection. Most alleles isolated in the Great Lakes region were closely related to the components of the trivalent Muguga vaccine. CONCLUSIONS: Our findings suggest that the extensive sequence diversity of T. parva and its biogeographical distribution mainly depend on host migration and agro-ecological conditions driving tick population dynamics. Such patterns are likely to contribute to the epidemic and unstable endemic situations of ECF in the region. However, the fact that ubiquitous alleles are genetically similar to the components of the Muguga vaccine together with the limited geographical clustering may justify testing the existing trivalent vaccine for cross-immunity in the region.
Asunto(s)
Variación Antigénica , Antígenos de Protozoos/genética , Theileria parva/genética , África Central , Antígenos de Protozoos/inmunología , Genotipo , Polimorfismo Genético , Análisis de Secuencia de ADN , Theileria parva/inmunologíaRESUMEN
The cattle tick Rhipicephalus microplus is the most important arthropod vector of livestock diseases globally. Since its introduction in West Africa a decade ago, it has been reported in Ivory Coast, Benin, Togo, Mali, Burkina Faso and Nigeria with potentially far-reaching adverse impacts on the livestock sector in the region. Cameroon is located on a major route for transboundary cattle trade between Central and West Africa and it is therefore at risk from R. microplus invasion. This study investigated the occurrence of R. microplus in Cameroon, the genetic polymorphism of the tick and population structure of isolates from different regions of the country to provide data that underpin the design of future vector control programs. A cross-sectional survey was conducted in which ticks were collected from cattle at 54 sites across the five Agroecological zones (AEZs) within Cameroon. Tick identity (sex and species) was assigned using taxonomic keys. Species identity was confirmed through amplification and sequencing of the mitochondrial COI and 16S rRNA genes. A total of 7091 ticks were collected out of which 1112 (15.6%) were morphologically identified as R. microplus. The presence of R. microplus was confirmed in 4 out of 5 agroecological zones. Only two haplotypes were identified by both COI and 16S rRNA genes, indicating a very low divergence in the genetic structure of the R. microplus population in Cameroon. 16S rRNA sequence analysis revealed a new haplotype specific to Cameroon. Phylogenetic trees revealed that all isolates of R. microplus from Cameroon were grouped into the previously described Africa/Americas clade. Application of a niche modelling algorithm to R. microplus distribution in Cameroon predicted that suitable habitat for the tick extended into southern Nigeria. This study demonstrated for the first time the presence of R. microplus in Cameroon. Genetic diversity tests indicate that the tick has not evolved significantly since the initial introduction to West Africa. We suggest further longitudinal studies to better define the spatial and temporal expansion of the range of the tick and the drivers of this spread.
Asunto(s)
Enfermedades de los Bovinos/epidemiología , Monitoreo Epidemiológico/veterinaria , Polimorfismo Genético , Rhipicephalus/genética , Infestaciones por Garrapatas/veterinaria , Algoritmos , Distribución Animal , Animales , Camerún/epidemiología , Bovinos , Enfermedades de los Bovinos/parasitología , Côte d'Ivoire/epidemiología , Estudios Transversales , Variación Genética , Haplotipos , Filogenia , ARN Ribosómico 16S/genética , Infestaciones por Garrapatas/epidemiologíaRESUMEN
BACKGROUND: The ixodid tick Rhipicephalus appendiculatus is the main vector of Theileria parva, wich causes the highly fatal cattle disease East Coast fever (ECF) in sub-Saharan Africa. Rhipicephalus appendiculatus populations differ in their ecology, diapause behaviour and vector competence. Thus, their expansion in new areas may change the genetic structure and consequently affect the vector-pathogen system and disease outcomes. In this study we investigated the genetic distribution of R. appendiculatus across agro-ecological zones (AEZs) in the African Great Lakes region to better understand the epidemiology of ECF and elucidate R. appendiculatus evolutionary history and biogeographical colonization in Africa. METHODS: Sequencing was performed on two mitochondrial genes (cox1 and 12S rRNA) of 218 ticks collected from cattle across six AEZs along an altitudinal gradient in the Democratic Republic of Congo, Rwanda, Burundi and Tanzania. Phylogenetic relationships between tick populations were determined and evolutionary population dynamics models were assessed by mismach distribution. RESULTS: Population genetic analysis yielded 22 cox1 and 9 12S haplotypes in a total of 209 and 126 nucleotide sequences, respectively. Phylogenetic algorithms grouped these haplotypes for both genes into two major clades (lineages A and B). We observed significant genetic variation segregating the two lineages and low structure among populations with high degree of migration. The observed high gene flow indicates population admixture between AEZs. However, reduced number of migrants was observed between lowlands and highlands. Mismatch analysis detected a signature of rapid demographic and range expansion of lineage A. The star-like pattern of isolated and published haplotypes indicates that the two lineages evolve independently and have been subjected to expansion across Africa. CONCLUSIONS: Two sympatric R. appendiculatus lineages occur in the Great Lakes region. Lineage A, the most diverse and ubiquitous, has experienced rapid population growth and range expansion in all AEZs probably through cattle movement, whereas lineage B, the less abundant, has probably established a founder population from recent colonization events and its occurrence decreases with altitude. These two lineages are sympatric in central and eastern Africa and allopatric in southern Africa. The observed colonization pattern may strongly affect the transmission system and may explain ECF endemic instability in the tick distribution fringes.
Asunto(s)
Vectores Arácnidos/genética , Variación Genética , Rhipicephalus/genética , Theileria parva/fisiología , Theileriosis/parasitología , África/epidemiología , Animales , Vectores Arácnidos/parasitología , Bovinos , Estructuras Genéticas , Genética de Población , Mitocondrias/genética , Filogenia , Filogeografía , Dinámica Poblacional , Rhipicephalus/parasitología , Theileriosis/epidemiologíaRESUMEN
OBJECTIVES: We evaluated the analytical performance of 24 immunoassays using the Beckman Coulter DxI 800 immunoassay systems at Mayo Clinic, Rochester, MN for trueness, precision, detection limits, linearity, and consistency (across instruments and reagent lots). METHODS: Clinically oriented performance goals were defined using the following methods: trueness-published desirable accuracy limits, precision-published desirable biologic variation; detection limits - 0.1 percentile of patient test values, linearity - 50% of total error, and consistency-percentage test values crossing key decision points. Local data were collected for precision, linearity, and consistency. Data were provided by Beckman Coulter, Inc. for trueness and detection limits. RESULTS: All evaluated assays except total thyroxine were within the proposed goals for trueness. Most of the assays met the proposed goals for precision (86% of intra-assay results and 75% of inter-assay results). Five assays had more than 15% of the test results below the minimum detection limits. Carcinoembryonic antigen, total thyroxine and free triiodothyronine exceeded the proposed goals of ±6.3%, ±5% and ±5.7% for dilution linearity. All evaluated assays were within the proposed goals for instrument consistency. Lot-to-lot consistency results for cortisol, ferritin and total thyroxine exceeded the proposed goals of 3.3%, 11.4% and 7% at one medical decision level, while vitamin B12 exceeded the proposed goals of 5.2% and 3.8% at two decision levels. CONCLUSIONS: The Beckman Coulter DxI 800 immunoassay system meets most of these proposed goals, even though these clinically focused performance goals represent relatively stringent limits.
Asunto(s)
Inmunoensayo/métodos , Ferritinas/análisis , Reproducibilidad de los Resultados , Tiroxina/análisis , Triyodotironina/análisisRESUMEN
Water buffalo industry has become a profitable activity worldwide, including the Northeast of Argentina (NEA). However, research on diseases affecting this species is scarce. The aim of the present study was to detect antibodies against Brucella abortus, Leptospira spp., Neospora caninum, Toxoplasma gondii, and Sarcocystis spp. in 500 water buffalo cows from five ranches (100 animals each) in the NEA. Serum samples were tested for B. abortus by fluorescence polarization assay, Leptospira spp. by microagglutination test, and N. caninum, T. gondii, and Sarcocystis spp. by indirect fluorescent antibody tests. Overall, the proportion of seropositive animals was 6.4, 22.2, 42.2, 25.4, and 50.8 % for brucellosis, leptospirosis, neosporosis, toxoplasmosis, and sarcocystosis, respectively. The proportion of seropositive animals for all diseases was statistically different among herds (p < 0.05). Statistical differences were also detected among age groups for brucellosis and neosporosis (p < 0.05). The detection of specific antibodies to B. abortus, Leptospira spp., and several Apicomplexa protozoans in water buffaloes in the NEA is reported in this study.
Asunto(s)
Búfalos/microbiología , Búfalos/parasitología , Leptospira/aislamiento & purificación , Neospora/aislamiento & purificación , Sarcocystis/aislamiento & purificación , Toxoplasma/aislamiento & purificación , Animales , Anticuerpos Antibacterianos/sangre , Anticuerpos Antiprotozoarios/sangre , Argentina/epidemiología , Infecciones Bacterianas/epidemiología , Infecciones Bacterianas/microbiología , Infecciones Bacterianas/veterinaria , Brucella abortus/aislamiento & purificación , Análisis Discriminante , Femenino , Infecciones Protozoarias en Animales/epidemiología , Infecciones Protozoarias en Animales/parasitología , Estudios SeroepidemiológicosRESUMEN
Several cell functions related to growth and survival regulation have been attributed specifically to the membrane form of heparin-binding EGF-like growth factor (proHB-EGF), rather than to the diffusible, processed HB-EGF isoform. These findings suggest the existence of a functional binding partner specifically for the membrane form of the growth factor. In this study we have identified the prosurvival cochaperone, BAG-1, as a protein that interacts with the cytoplasmic tail domain of proHB-EGF. Interaction between BAG-1 and the 24-amino acid proHB-EGF cytoplasmic tail was initially identified in a yeast two-hybrid screen and was confirmed in mammalian cells. The proHB-EGF tail bound BAG-1 in an hsp70-independent manner and within a 97-amino acid segment that includes the ubiquitin homology domain in BAG-1 but does not include the hsp70 binding site. Effects of BAG-1 and proHB-EGF co-expression were demonstrated in cell adhesion and cell survival assays and in quantitative assays of regulated secretion of soluble HB-EGF. Because the BAG-1 binding site is not present on the mature, diffusible form of the growth factor, these findings suggest a new mechanism by which proHB-EGF, in isolation from the diffusible form, can mediate cell signaling events. In addition, because effects of BAG-1 on regulated secretion of soluble HB-EGF were also identified, this interaction has the potential to alter the signaling capabilities of both the membrane-anchored and the diffusible forms of the growth factor.
Asunto(s)
Proteínas Portadoras/química , Proteínas Portadoras/metabolismo , Membrana Celular/metabolismo , Citoplasma/metabolismo , Factor de Crecimiento Epidérmico/metabolismo , Heparina/metabolismo , Animales , Apoptosis , Sitios de Unión , Células CHO , Células COS , Adhesión Celular/efectos de los fármacos , División Celular , Supervivencia Celular , Cricetinae , Proteínas de Unión al ADN , Relación Dosis-Respuesta a Droga , Etopósido/farmacología , Glutatión Transferasa/metabolismo , Proteínas HSP70 de Choque Térmico/metabolismo , Humanos , Microscopía Confocal , Inhibidores de la Síntesis del Ácido Nucleico/farmacología , Unión Proteica , Isoformas de Proteínas , Estructura Terciaria de Proteína , Proteínas Recombinantes de Fusión/metabolismo , Factores de Tiempo , Factores de Transcripción , Transfección , Células Tumorales Cultivadas , Técnicas del Sistema de Dos Híbridos , Ubiquitinas/metabolismoRESUMEN
Antecedentes: El cisatracurio, un nuevo relajante muscular de acción intermedia, reúne varios de los criterios que hacen al "relajante muscular ideal", salvo su tiempo de latencia de 3 a 5 minutos con dosis habituales de intubación. Objetivos: 1.- acortar el tiempo de latencia administrando 6 dosis efectivas 95 (6 DE 95); 2.- comprobar parámetros de estabilidad cardiovascular y ausencia de signos clínicos de liberación de histamina y 3.- obtener una prolongación de la duración clínica no mayor del 7 por ciento. Lugar de aplicación: Hospital Gral. de Agudos Dr. J. M. Ramos Mejía, Buenos Aires, Argentina, enero-abril de 1999. Diseño: Ensayo clínico prospectivo aleatorizado doble ciego con observador único. Población: 40 pacientes para cirugía electiva, ASA I-II, 18-70 años. Método: Los 40 pacientes fueron asignados aleatoriamente a dos grupos: M (0,3 mg/kg de cisatracurio) y C (grupo control, 0,1 mg/kg de cisatracurio). Se registraron los valores de tensión arterial sistólica, tensión arterial diastólica y frecuencia cardíaca no invasivas, oximetría de pulso, trazado electrocardioscópico y monitoreo de la función neuromuscular por acelerometría. La anestesia fue estandarizada : premedicación con fentanilo y metoclopramida, inducción con propofol y mantenimiento con isofluorano en mezcla de óxido nitroso y oxígeno. Para el análisis estadístico se empleó el Test de t a dos colas para dos muestras de observaciones independientes, considerando estadísticamente significativo un valor de P < 0,05. Resultados: El tiempo de latencia fue de 76,3 ñ 13,69 seg y la duración clínica, de 75,4 ñ 7,14 min en el grupo M, y de 210 ñ 30,47 seg y 47,4 ñ 8,17 min, respectivamente, en el Grupo C, siendo estas diferencias estadísticamente significativas para P < 0,01. La duración clínica se prolongó en un 59 por ciento respecto del grupo control. No hubo diferencias significativas en los valores de tensión arterial y frecuencia cardíaca. No hubo manifestaciones clínicas de liberación de histamina. Conclusiones: La administración de 6 DE 95 de cisatracurio es una técnica segura y útil frente al rocuronio y a la succinilcolina para lograr un corto tiempo de latencia para la intubación traqueal. (AU)
Asunto(s)
Humanos , Masculino , Femenino , Adulto , Persona de Mediana Edad , Anciano , Relajantes Musculares Centrales/farmacocinética , Tiempo de Reacción/efectos de los fármacos , Atracurio/administración & dosificación , Sistema Cardiovascular/efectos de los fármacos , Monitoreo Fisiológico , Intubación Intratraqueal , Hemodinámica/efectos de los fármacosRESUMEN
Antecedentes: El cisatracurio, un nuevo relajante muscular de acción intermedia, reúne varios de los criterios que hacen al "relajante muscular ideal", salvo su tiempo de latencia de 3 a 5 minutos con dosis habituales de intubación. Objetivos: 1.- acortar el tiempo de latencia administrando 6 dosis efectivas 95 (6 DE 95); 2.- comprobar parámetros de estabilidad cardiovascular y ausencia de signos clínicos de liberación de histamina y 3.- obtener una prolongación de la duración clínica no mayor del 7 por ciento. Lugar de aplicación: Hospital Gral. de Agudos Dr. J. M. Ramos Mejía, Buenos Aires, Argentina, enero-abril de 1999. Diseño: Ensayo clínico prospectivo aleatorizado doble ciego con observador único. Población: 40 pacientes para cirugía electiva, ASA I-II, 18-70 años. Método: Los 40 pacientes fueron asignados aleatoriamente a dos grupos: M (0,3 mg/kg de cisatracurio) y C (grupo control, 0,1 mg/kg de cisatracurio). Se registraron los valores de tensión arterial sistólica, tensión arterial diastólica y frecuencia cardíaca no invasivas, oximetría de pulso, trazado electrocardioscópico y monitoreo de la función neuromuscular por acelerometría. La anestesia fue estandarizada : premedicación con fentanilo y metoclopramida, inducción con propofol y mantenimiento con isofluorano en mezcla de óxido nitroso y oxígeno. Para el análisis estadístico se empleó el Test de t a dos colas para dos muestras de observaciones independientes, considerando estadísticamente significativo un valor de P < 0,05. Resultados: El tiempo de latencia fue de 76,3 ñ 13,69 seg y la duración clínica, de 75,4 ñ 7,14 min en el grupo M, y de 210 ñ 30,47 seg y 47,4 ñ 8,17 min, respectivamente, en el Grupo C, siendo estas diferencias estadísticamente significativas para P < 0,01. La duración clínica se prolongó en un 59 por ciento respecto del grupo control. No hubo diferencias significativas en los valores de tensión arterial y frecuencia cardíaca. No hubo manifestaciones clínicas de liberación de histamina. Conclusiones: La administración de 6 DE 95 de cisatracurio es una técnica segura y útil frente al rocuronio y a la succinilcolina para lograr un corto tiempo de latencia para la intubación traqueal.
Asunto(s)
Humanos , Masculino , Femenino , Adulto , Persona de Mediana Edad , Atracurio/administración & dosificación , Sistema Cardiovascular/efectos de los fármacos , Relajantes Musculares Centrales/farmacocinética , Tiempo de Reacción , Hemodinámica , Intubación Intratraqueal , Monitoreo FisiológicoRESUMEN
By the year 2005, the Human Genome Project is expected to have mapped and sequenced all of the estimated 100,000 genes that encode the various proteins found in human cells. Defining the role of each gene, and using that information to redirect its action when therapeutic intervention is required, is one of the major assignments for molecular medicine and molecular urology in the post-genome era. This challenge to determine gene function, and to do it cost-effectively and on a large scale, has driven development of new technologies that can more efficiently flag genes that are likely targets for therapeutic intervention. Yeast-based genetic assays that detect protein-protein interactions in vivo offer many of the features required of a practical "gene-flagging" strategy for identifying genes that might be functionally manipulated to achieve therapeutic goals. In the past few years, the yeast-based assays collectively referred to as "two-hybrid interaction traps" or simply "two-hybrid systems" have become increasingly important tools for experimental analysis of gene function. This review presents an overview of the principles of yeast-based two-hybrid analysis, examines some specific applications of the technique of interest to urologic investigators, and discusses some key points that a basic urologic investigator new to the technology would want to consider when designing or evaluating a yeast-based two-hybrid project.
Asunto(s)
Urología/métodos , Levaduras/genética , Genoma Humano , Humanos , Hibridación GenéticaRESUMEN
Partial forced expiratory flow-volume curves obtained by the rapid chest compression technique are being widely used to assess pulmonary function in infants and young children. The aim of this study is to assess whether in this age group flow limitation is achieved with the partial forced expiratory flow-volume curve with rapid chest compression. In eight infants and young children sedated with chloral hydrate, flow-volume curves were obtained by regular rapid chest compression technique, end-inspiratory airway occlusion prior to rapid chest compression, and expiratory clamping prior to rapid chest compression. In each technique, beginning with a cuff pressure of 20 cmH2O, the cuff pressure was increased by 10 cmH2O increments until the compression pressure reached 90 cmH2O. Maximal flow-volume curves were generated by each technique. End-inspiratory occlusion prior to rapid chest compression caused higher flows over the entire phase of expiration than the regular rapid chest compression. This increase could be observed over the entire phase of expiration. Forced expiratory flow at 50% and at 75% of vital capacity (V50 and V75) with regular rapid chest compression were 207 +/- 44 ml.s-1 (mean +/- SD) and 138 +/- 59 ml.sec-1, respectively. When end-inspiratory occlusion preceded rapid chest compression, V50 and V75 increased to 283 +/- 114 and 206 +/- 61 ml.sec-1 respectively, with a mean increase in V50 of 34% and in V75 of 31%. When expiratory clamping preceded the compression, even higher expiratory flows resulted.(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Pruebas de Función Respiratoria , Preescolar , Humanos , Lactante , Mediciones del Volumen Pulmonar , Flujo Espiratorio Máximo , Pruebas de Función Respiratoria/métodosRESUMEN
Glutamate receptors (GluRs) mediate excitatory neurotransmission and may have important roles in central nervous system disorders. To characterize the human GLUR5 gene, which is located on human chromosome 21q22.1, we isolated cDNAs, genomic phage lambda clones, and yeast artificial chromosomes (YACs) and developed sequence tagged sites (STSs) and simple sequence length polymorphisms (SSLPs) for GLUR5. Genetic mapping with a tetranucleotide AGAT repeat named GLUR5/AGAT (six alleles observed, 70% heterozygosity) placed GLUR5 5 cM telomeric to APP (D21S210) and 3 cM centromeric to SOD1 (D21S223). The human GLUR5 gene is located near the familial amyotrophic lateral sclerosis (FALS) locus; linkage analysis of GLUR5 SSLPs in FALS pedigrees yielded negative lod scores, consistent with the recent association of the FALS locus with the SOD1 gene. Physical mapping of GLUR5 using a YAC contig suggested that the GLUR5 gene spans approximately 400-500kb, and is within 280kb of D21S213. The large size of the GLUR5 gene raises questions regarding its functional significance. Our GLUR5 YAC contig includes clones found in the Genethon chromosome 21 YAC contig, and reference to the larger contig indicates the orientation centromere--D21S213-GLUR5 5' end-GLUR5/AGAT--GLUR5 3' end--SOD1. The development of GLUR5/AGAT should permit rapid determination of the status of the GLUR5 gene in individuals with partial trisomy or monosomy of chromosome 21. Such studies may provide insights concerning the possible role of GLUR5 in Down syndrome.
Asunto(s)
Mapeo Cromosómico , Cromosomas Humanos Par 21 , Receptores de Glutamato/genética , Secuencia de Bases , Cromosomas Artificiales de Levadura , Clonación Molecular , Humanos , Escala de Lod , Datos de Secuencia Molecular , Polimorfismo Genético , Recombinación Genética , Secuencias Repetitivas de Ácidos Nucleicos/genética , Análisis de Secuencia de ADN , Lugares Marcados de SecuenciaRESUMEN
Potent and highly selective adenosine A1 and A2 receptor agonists were bilaterally injected into the nucleus accumbens of mice 10 min prior to inhibitory avoidance training. Retention of the inhibitory avoidance response was assessed 24 h after training. Intra-ACB activation of A1 receptors, but not A2a receptor activation, significantly impaired the performance of mice during the subsequent retention test. Furthermore, the retention deficit produced by activation of A1 receptors was significantly attenuated by pretreating mice with a highly selective A1 receptor antagonist. These findings suggest that endogenous adenosine may modulate information processing in the ventral striatum via adenosine A1 receptors.
Asunto(s)
Reacción de Prevención/fisiología , Inhibición Neural/fisiología , Núcleo Accumbens/fisiología , Receptores Purinérgicos P1/fisiología , Adenosina/análogos & derivados , Adenosina/farmacología , Animales , Reacción de Prevención/efectos de los fármacos , Mapeo Encefálico , Electrochoque , Miedo/efectos de los fármacos , Miedo/fisiología , Masculino , Recuerdo Mental/efectos de los fármacos , Recuerdo Mental/fisiología , Ratones , Ratones Endogámicos ICR , Inhibición Neural/efectos de los fármacos , Núcleo Accumbens/efectos de los fármacos , Fenetilaminas/farmacología , Receptores Purinérgicos P1/clasificación , Receptores Purinérgicos P1/efectos de los fármacosRESUMEN
An unusual strain of Pseudomonas putida UUC-1 capable of growth at high salicylate concentration (10 gl-1) was investigated with the aim of developing an assay and a biosensor system for determining salicylate in body fluids by utilizing the salicylate hydroxylase enzyme. Medium and growth condition optimization were carried out under chemostat conditions. The highest biomass yield was at 4.0 gl-1 salicylate, 25 degrees C, pH 6.5, and 0.2 h-1 dilution rate. Growth occurred at up to 0.45 h-1 dilution rate, producing 236 Ul-1 enzyme activity and an output of 424 U h-1. The activity and productivity were higher than any reported in the literature for this enzyme. It had a Km value of 2.07 +/- 0.32 microM and an M(r) of approximately 43,000. In addition, its specific activity in the crude extract (0.8-0.9 U mg-1 protein) was similar to the commercially available enzyme. No plasmid DNA was detected in this strain, and no salicylate-negative isolates were obtained when curing with mitomycin C. It is therefore proposed that our strain has a chromosomally located inducible salicylate hydroxylase gene that enables it to grow at high salicylate. This strain also offers a means of cheaply producing large quantities of the enzyme through standard fermentation techniques.
Asunto(s)
Oxigenasas de Función Mixta/biosíntesis , Pseudomonas putida/enzimología , Técnicas Biosensibles , Biotecnología , Medios de Cultivo , Fermentación , Oxigenasas de Función Mixta/aislamiento & purificación , Plásmidos/aislamiento & purificación , Pseudomonas putida/genética , Pseudomonas putida/crecimiento & desarrollo , Salicilatos/metabolismo , Ácido SalicílicoRESUMEN
We have isolated and sequenced cDNAs representing the full-length (2987-bp) gene for dihydrolipoyl succinyltransferase (E2k component) of the human alpha-ketoglutarate dehydrogenase complex (KGDHC) from a human fetal brain cDNA library. The E2k cDNA was mapped to human chromosome 14 using a somatic cell hybrid panel, and more precisely to band 14q24.3 by in situ hybridization. This cDNA also cross-hybridized to an apparent E2k pseudogene on chromosome 1p31. Northern analysis revealed the E2k gene to be ubiquitously expressed in peripheral tissues and brain. Interestingly, chromosome 14q24.3 has recently been reported to contain gene defects for an early-onset form of familial Alzheimer's disease and for Machado-Joseph disease. Future studies will be necessary to determine whether the E2k gene plays a role in either of these two disorders.
Asunto(s)
Aciltransferasas/genética , Secuencia de Aminoácidos , Secuencia de Bases , Northern Blotting , Southern Blotting , Encéfalo/enzimología , Mapeo Cromosómico , Cromosomas Humanos Par 1 , Cromosomas Humanos Par 14 , Clonación Molecular , ADN Complementario , Humanos , Hibridación in Situ , Datos de Secuencia Molecular , SeudogenesRESUMEN
Inherited Alzheimer's disease is a genetically heterogeneous disorder that involves gene defects on at least five chromosomal loci. Three of these loci have been found by genetic linkage studies to reside on chromosomes 21, 19, and 14. On chromosomes 21, the gene encoding the precursor protein of Alzheimer-associated amyloid (APP) has been shown to contain several mutations in exons 16 and 17 which account for roughly 2-3% of familial Alzheimer's disease (FAD). The other loci include what appears to be a susceptibility gene on chromosome 19 associated with late-onset (> 65 years) FAD, and a major early-onset FAD gene defect on the long arm of chromosome 14. In other early- and late-onset FAD kindreds, the gene defects involved do not appear to be linked to any of these three loci, indicating the existence of additional and as of yet unlocalized FAD genes. This review provides a historical perspective of the search for FAD gene defects and summarizes the progress made in world-wide attempts to isolate and characterize the genes responsible for this disorder.
Asunto(s)
Enfermedad de Alzheimer/genética , Precursor de Proteína beta-Amiloide/genética , Genes , Mutación/genética , Enfermedad de Alzheimer/etiología , Cromosomas Humanos , Ligamiento Genético , Humanos , Modelos BiológicosRESUMEN
Hypoglycemia in patients with diabetes was treated by a standardized protocol in a 300-bed community hospital. Patient outcomes were studied retrospectively using data from medical records. Hypoglycemia was defined as a blood glucose level below 80 mg/dL when symptoms were present, or 60 mg/dL regardless of symptoms. Treatment consisted of 20 grams of fast-acting carbohydrate repeated every 15 minutes until the blood glucose level was above 80 mg/dL. The blood glucose level was then rechecked one hour after treatment. Data from 179 episodes were evaluated. In 153 (85%) cases, the blood glucose levels were within the target range (80-200 mg/dL) at the conclusion of treatment. In 135 (76%) cases, the blood glucose level at one hour following treatment was within the same target range.
Asunto(s)
Protocolos Clínicos/normas , Complicaciones de la Diabetes , Carbohidratos de la Dieta/administración & dosificación , Hipoglucemia/dietoterapia , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Hospitales Comunitarios , Humanos , Hipoglucemia/sangre , Hipoglucemia/etiología , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Resultado del TratamientoRESUMEN
Sequence tagged sites (STSs) have been proposed as a "common language" for comparing physical and genetic maps of the human genome produced by a variety of techniques. We have produced 44 STSs from 38 mapped loci on human chromosome 21. The STSs represent most of the loci designated as genetic reference or ordered physical framework markers, along with a number of others chosen to span all regions of 21q. Of the STSs, 12 are from gene segments, including 4 from exons of the APP gene encoding the amyloid beta protein precursor, and 32 mark anonymous DNA loci. These STSs make each of the corresponding loci readily accessible to the research community without the need for exchange of clones. These sites also represent multiple start points for the isolation of YAC clones that should permit overlapping the entire chromosome 21 long arm as cloned DNA.
Asunto(s)
Cromosomas Humanos Par 21 , Marcadores Genéticos , Lugares Marcados de Secuencia , Animales , Secuencia de Bases , Mapeo Cromosómico , ADN de Cadena Simple , Humanos , Células Híbridas , Ratones , Datos de Secuencia Molecular , Reacción en Cadena de la PolimerasaRESUMEN
This controlled experimental study examines the effect of two teaching methods on achievement outcomes from a 15-week, 2 credit hour semester course taught at two midwestern universities. Students were randomly assigned to either computer-managed instruction in which faculty function as tutors or the traditional classroom course of study. In addition, the effects of age, grade point average, attitudes toward computers, and satisfaction with the course on teaching method were analyzed using analysis of covariance. Younger students achieved better scores than did older students. Regardless of teaching method, however, neither method appeared to be better than the other for teaching course content. Students did not prefer one method over the other as indicated by their satisfaction scores. With demands upon university faculty to conduct research and publish, alternative methods of teaching that free faculty from the classroom should be considered. This study suggests that educators can select such an alternative teaching method to traditional classroom teaching without sacrificing quality education for certain courses.