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1.
J Perinatol ; 37(4): 436-440, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-27977019

RESUMO

OBJECTIVE: Small-for-gestational-age (SGA) neonates, infants of diabetic mothers (IDM) and very-low-birth weight premature neonates (VLBW) are reported to have increased risk for developing iron deficiency and possibly associated neurocognitive delays. STUDY DESIGN: We conducted a pilot study to assess iron status at birth in at-risk neonates by measuring iron parameters in umbilical cord blood from SGA, IDM, VLBW and comparison neonates. RESULTS: Six of the 50 infants studied had biochemical evidence of iron deficiency at birth. Laboratory findings consistent with iron deficiency were found in one SGA, one IDM, three VLBW, and one comparison infant. None of the infants had evidence of iron deficiency anemia. CONCLUSIONS: Evidence of biochemical iron deficiency at birth was found in 17% of screened neonates. Studies are needed to determine whether these infants are at risk for developing iron-limited erythropoiesis, iron deficiency anemia or iron-deficient neurocognitive delay.


Assuntos
Anemia Ferropriva/sangue , Recém-Nascido Pequeno para a Idade Gestacional/sangue , Recém-Nascido de muito Baixo Peso/sangue , Ferro/sangue , Estudos de Casos e Controles , Diabetes Gestacional , Feminino , Ferritinas/sangue , Sangue Fetal/química , Humanos , Recém-Nascido , Modelos Lineares , Masculino , Projetos Piloto , Gravidez , Gravidez em Diabéticas , Estudos Prospectivos , Fatores de Risco , Utah
2.
Br J Cancer ; 108(2): 450-60, 2013 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-23299538

RESUMO

BACKGROUND: When designing therapeutic short-interfering RNAs (siRNAs), off-target effects (OTEs) are usually predicted by computational quantification of messenger RNAs (mRNAs) that contain matches to the siRNA seed sequence in their 3' UTRs. It is assumed that the higher the number of predicted transcriptional OTEs, the greater the size of the actual OTE signature and the more detrimental the phenotypic consequences in target-negative cells. METHODS: We tested this general assumption by investigating the OTEs of potential therapeutic siRNAs targeting the human papillomavirus (HPV) type-16 E7 oncogene. We studied HPV-negative squamous epithelial cells, from normal cervix (NCx) and skin (HaCaT), which would be vulnerable to 'bystander' OTEs following transfection in vivo. RESULTS: We observed no correlation between the number of computationally predicted OTEs and the actual number of seed-dependent OTEs (P=0.76). On average only 20.5% of actual transcriptional OTEs were seed-dependent (i.e., predicted). The unpredicted OTEs included stimulation of innate immune pathways, as well as indirect (downstream) effects of other OTEs, which affected important cancer-associated pathways. Although most significant OTEs observed were seen in both NCx and HaCaT cells, only 0-5.9% of differentially expressed genes overlapped between the two cell types. CONCLUSION: These data do not support the assumption that actual OTEs correlate well with predicted OTEs.


Assuntos
Papillomavirus Humano 16/genética , Proteínas E7 de Papillomavirus/genética , Neoplasias do Colo do Útero/virologia , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/virologia , Linhagem Celular Tumoral , Colo do Útero/citologia , Células Epiteliais/virologia , Feminino , Humanos , Interferência de RNA , RNA Interferente Pequeno , Pele/citologia , Neoplasias do Colo do Útero/genética
3.
Heredity (Edinb) ; 100(2): 207-19, 2008 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-17551524

RESUMO

A microbial species concept is crucial for interpreting the variation detected by genomics and environmental genomics among cultivated microorganisms and within natural microbial populations. Comparative genomic analyses of prokaryotic species as they are presently described and named have led to the provocative idea that prokaryotes may not form species as we think about them for plants and animals. There are good reasons to doubt whether presently recognized prokaryotic species are truly species. To achieve a better understanding of microbial species, we believe it is necessary to (i) re-evaluate traditional approaches in light of evolutionary and ecological theory, (ii) consider that different microbial species may have evolved in different ways and (iii) integrate genomic, metagenomic and genome-wide expression approaches with ecological and evolutionary theory. Here, we outline how we are using genomic methods to (i) identify ecologically distinct populations (ecotypes) predicted by theory to be species-like fundamental units of microbial communities, and (ii) test their species-like character through in situ distribution and gene expression studies. By comparing metagenomic sequences obtained from well-studied hot spring cyanobacterial mats with genomic sequences of two cultivated cyanobacterial ecotypes, closely related to predominant native populations, we can conduct in situ population genetics studies that identify putative ecotypes and functional genes that determine the ecotypes' ecological distinctness. If individuals within microbial communities are found to be grouped into ecologically distinct, species-like populations, knowing about such populations should guide us to a better understanding of how genomic variation is linked to community function.


Assuntos
Cianobactérias/classificação , Cianobactérias/genética , Ecossistema , Genômica , Meio Ambiente , Genética Populacional
4.
J Anim Ecol ; 76(1): 135-8, 2007 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17184361

RESUMO

1. Empirical studies show that average growth of stream-dwelling salmon and trout often declines with increasing density in a characteristic concave relationship. However, the mechanisms that generate negative density-growth relationships in populations in natural streams are not certain. 2. In a recent study, Imre, Grant & Cunjak (2005; Journal of Animal Ecology, 74, 508-516) argue that density-dependent growth due to exploitative competition for prey causes the negative density-growth relationships for stream salmonids. They argue that the concave shape of empirical density-growth relationships is consistent with a simple model of exploitative competition and not consistent with interference competition for space. 3. We use a simple model to show that competition for space can yield concave density-growth relationships consistent with the empirical pattern when individuals compete for foraging sites that vary spatially in quality and lower-quality sites predominate. Thus, the predictions of the exploitative competition and spatial competition models overlap. 4. The shape of the density-growth relationship does not differentiate between candidate mechanisms underlying density-dependent growth for stream salmonids. Our results highlight the general problem with determining the mechanism driving an ecological process from patterns in observational data within the context of linking population demographics to habitat structure and animal behaviour.


Assuntos
Comportamento Competitivo/fisiologia , Ecossistema , Comportamento Alimentar/fisiologia , Salmonidae/crescimento & desenvolvimento , Animais , Modelos Biológicos , Densidade Demográfica , Rios
5.
Cytotherapy ; 6(4): 318-27, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-16146884

RESUMO

BACKGROUND: The adequacy of HPC collection for BMT is typically assessed by the number of CD34 cells. However, during a series of leukapheresis procedures (LP) the CD34 value on the final HPC product may not be available for testing until late evening, sometimes resulting in additional, retrospectively unnecessary, LP in order to ensure an adequate HPC collection (>5x10(6) CD34/kg). We hypothesized that an estimate of the CD34 content of HPC products prior to 16:00 h on the day of LP would permit improved HPC collection planning. We therefore assessed the effectiveness of predicting the total amount of CD34 cells that would be collected in a given LP by either (a) the concentration of CD34 cells/microL in peripheral blood prior to LP (pre-CD34) or (b) the predicted total amount of CD34 cells to be collected based on sampling the LP product at the mid-point of each LP. We also compared the number of LP per patient and total HPC collected for the study group with data from the previous calendar year. METHODS: Allogeneic and autologous BMT donors who completed a 20-L HPC collection between September 2002 and February 2003 were eligible. CD34 cells were measured on blood drawn prior to LP and from the HPC product at the mid-point (10 L) of LP. The CD34 content of the final LP was predicted by doubling the value of total CD34 cells at the mid-run (MRp-CD34). The MRp-CD34/kg and the cumulative CD34/kg collected were made available before 16:00 h and used to determine the need for additional LP. The true CD34 content of each HPC collection was also measured from the final product the next day (CD34-FP). RESULTS: A 20-L LP was completed and data were available from 31 patients and nine allogeneic donors who underwent a total of 85 LP for diagnoses, including 11 myeloma, 10 lymphoma, seven HD, three acute leukemia and five others. The mean (range) and correlation (R2) vs. the CD34-FP were, for pre-CD34, 54 CD34/microL (0.3-232), R2=0.66 (P<0.01), and for MRp-CD34, 3.2x10(6) CD34/kg (0.04-22.48), R2=0.90 (P<0.01). The mean number of CD34/kg collected per LP in the patients/donors was 3.4x10(6) CD34/kg (0.05-18.94). The median number of CD34 cells employed for transplant in the study group vs. controls (5.7 vs. 5.6x10(6)/kg) and the time to engraftment of neutrophils (12 vs. 11 days) and platelets (12 vs. 12 days) was similar to historical controls. However, the study group had a significantly lower median number of LP (three vs. two; P<0.02) to obtain the required collection of 5x10(6) CD34 cells/kg. DISCUSSION: Both the pre-CD34 and the MRp-CD34 were significantly correlated with CD34-FP. However, the CD34-FP was more reliably predicted by MRp-CD34. Early availability of mid-run CD34 values was associated with a significant reduction in the number of LP required to collect 5x10(6) CD34 cells/kg, without reduction in the number of CD34 cells for transplant or prolongation of days to neutrophil or platelet engraftment.


Assuntos
Antígenos CD34/metabolismo , Transplante de Medula Óssea , Mobilização de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/citologia , Leucaférese , Células-Tronco Hematopoéticas/metabolismo , Humanos
6.
J Biol Chem ; 276(46): 43049-55, 2001 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-11562378

RESUMO

Saccharomyces cerevisiae can accumulate iron through the uptake of siderophore-iron. Siderophore-iron uptake can occur through the reduction of the complex and the subsequent uptake of iron by the high affinity iron transporter Fet3p/Ftr1p. Alternatively, specific siderophore transporters can take up the siderophore-iron complex. The pathogenic fungus Candida albicans can also take up siderophore-iron. Here we identify a C. albicans siderophore transporter, CaArn1p, and characterize its activity. CaARN1 is transcriptionally regulated in response to iron. Through expression studies in S. cerevisiae strains lacking endogenous siderophore transporters, we demonstrate that CaArn1p specifically mediates the uptake of ferrichrome-iron. Iron-ferrichrome and gallium-ferrichrome, but not desferri-ferrichrome, could competitively inhibit the uptake of iron from ferrichrome. Uptake of siderophore-iron resulting from expression of CaARN1 under the control of the MET25-promoter in S. cerevisiae was independent of the iron status of the cells and of Aft1p, the iron-sensing transcription factor. These studies demonstrate that the expression of CaArn1p is both necessary and sufficient for the nonreductive uptake of ferrichrome-iron and suggests that the transporter may be the only required component of the siderophore uptake system that is regulated by iron and Aft1p.


Assuntos
Candida albicans/química , Candida albicans/metabolismo , Proteínas de Transporte/química , Proteínas de Transporte/fisiologia , Proteínas de Transporte de Cátions , Ferricromo/metabolismo , Proteínas Fúngicas , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/metabolismo , Sequência de Aminoácidos , Transporte Biológico , Northern Blotting , Western Blotting , Proteínas de Transporte/biossíntese , Relação Dose-Resposta a Droga , Ferro/metabolismo , Ferro/farmacocinética , Ferro/farmacologia , Cinética , Metionina/metabolismo , Dados de Sequência Molecular , Plasmídeos/metabolismo , Regiões Promotoras Genéticas , Homologia de Sequência de Aminoácidos , Sideróforos/farmacocinética
7.
Appl Environ Microbiol ; 67(9): 4365-71, 2001 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-11526049

RESUMO

Filamentous bacteria containing bacteriochlorophylls c and a were enriched from hypersaline microbial mats. Based on phylogenetic analyses of 16S rRNA gene sequences, these organisms form a previously undescribed lineage distantly related to Chloroflexus spp. We developed and tested a set of PCR primers for the specific amplification of 16S rRNA genes from filamentous phototrophic bacteria within the kingdom of "green nonsulfur bacteria." PCR products recovered from microbial mats in a saltern in Guerrero Negro, Mexico, were subjected to cloning or denaturing gradient gel electrophoresis and then sequenced. We found evidence of a high diversity of bacteria related to Chloroflexus which exhibit different distributions along a gradient of salinity from 5.5 to 16%.


Assuntos
Chlorobi/classificação , Chlorobi/genética , Ecossistema , Cloreto de Sódio , Microbiologia da Água , Chlorobi/crescimento & desenvolvimento , Meios de Cultura , Primers do DNA , Genes de RNAr , Dados de Sequência Molecular , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
8.
J Biol Chem ; 276(24): 10971-6, 2001 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-11439929

RESUMO

To assess the effects related to known and proposed biosynthetic pathways on the (13)C content of lipids and storage products of the photoautotrophic bacterium Chloroflexus aurantiacus, the isotopic compositions of bulk cell material, alkyl and isoprenoid lipids, and storage products such as glycogen and polyhydroxyalkanoic acids have been investigated. The bulk cell material was 13 per thousand depleted in (13)C relative to the dissolved inorganic carbon. Evidently, inorganic carbon fixation by the main carboxylating enzymes used by C. aurantiacus, which are assumed to use bicarbonate rather than CO(2), results in a relatively small carbon isotopic fractionation compared with CO(2) fixation by the Calvin cycle. Even carbon numbered fatty acids, odd carbon numbered fatty acids, and isoprenoid lipids were 14, 15, and 17-18 per thousand depleted in (13)C relative to the carbon source, respectively. Based on the (13)C contents of alkyl and isoprenoid lipids, a 40 per thousand difference in (13)C content between the carboxyl and methyl carbon from acetyl-coenzyme A has been calculated. Both sugars and polyhydroxyalkanoic acid were enriched in (13)C relative to the alkyl and isoprenoid lipids. To the best of our knowledge this is the first report in which the stable carbon isotopic composition of a large range of biosynthetic products in a photoautotrophic organism has been investigated and interpreted based on previously proposed inorganic carbon fixation and biosynthetic pathways. Our results indicate that compound-specific stable carbon isotope analysis may provide a rapid screening tool for carbon fixation pathways.


Assuntos
Alcanos/metabolismo , Carbono/metabolismo , Chlorobi/metabolismo , Glicogênio/metabolismo , Metabolismo dos Lipídeos , Isótopos de Carbono
9.
J Biol Chem ; 276(14): 10971-6, 2001 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-11145961

RESUMO

To assess the effects related to known and proposed biosynthetic pathways on the (13)C content of lipids and storage products of the photoautotrophic bacterium Chloroflexus aurantiacus, the isotopic compositions of bulk cell material, alkyl and isoprenoid lipids, and storage products such as glycogen and polyhydroxyalkanoic acids have been investigated. The bulk cell material was 13 per thousand depleted in (13)C relative to the dissolved inorganic carbon. Evidently, inorganic carbon fixation by the main carboxylating enzymes used by C. aurantiacus, which are assumed to use bicarbonate rather than CO(2), results in a relatively small carbon isotopic fractionation compared with CO(2) fixation by the Calvin cycle. Even carbon numbered fatty acids, odd carbon numbered fatty acids, and isoprenoid lipids were 14, 15, and 17-18 per thousand depleted in (13)C relative to the carbon source, respectively. Based on the (13)C contents of alkyl and isoprenoid lipids, a 40 per thousand difference in (13)C content between the carboxyl and methyl carbon from acetyl-coenzyme A has been calculated. Both sugars and polyhydroxyalkanoic acid were enriched in (13)C relative to the alkyl and isoprenoid lipids. To the best of our knowledge this is the first report in which the stable carbon isotopic composition of a large range of biosynthetic products in a photoautotrophic organism has been investigated and interpreted based on previously proposed inorganic carbon fixation and biosynthetic pathways. Our results indicate that compound-specific stable carbon isotope analysis may provide a rapid screening tool for carbon fixation pathways.


Assuntos
Alcanos/metabolismo , Carbono/metabolismo , Chlorobi/metabolismo , Glicogênio/metabolismo , Metabolismo dos Lipídeos , Biodegradação Ambiental , Radioisótopos de Carbono
10.
J Cell Sci ; 113 ( Pt 22): 4077-86, 2000 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-11058094

RESUMO

AP-3 is a heterotetrameric protein complex involved in intracellular vesicle transport. Molecular analyses show that Ap3b1, which encodes the AP-3 (&bgr;)3A subunit, is altered in pearl mice. To provide genetic evidence that mutation of Ap3b1 is responsible for the pearl phenotype and to determine the null phenotype, the Ap3b1 gene was disrupted by homologous recombination. Mice homozygous for the resulting allele, Ap3b1(LN), or compound heterozygotes with pearl, displayed phenotypes similar to those of pearl mice, confirming that Ap3b1 is the causal gene for pearl. Moreover, pearl is likely to be a hypomorph as the Ap3b1(LN) homozygotes had a lighter coat color and accumulated fewer of the micro3 and (&dgr;)3 subunits of AP-3 than did pearl mice. Finally, immunofluorescence analysis of fibroblasts and melanocytes cultured from Ap3b1(LN) homozygotes revealed that the lysosomal membrane proteins Lamp I and Lamp II and the melanosomal membrane protein tyrosinase were mislocalized. In particular, the Lamp proteins were clustered on the cell surface. These findings strengthen the evidence for an alternate pathway via the plasma membrane for cargo normally transported to organelles by AP-3.


Assuntos
Complexo 3 de Proteínas Adaptadoras , Antígenos CD/metabolismo , Proteínas Heterotriméricas de Ligação ao GTP/metabolismo , Glicoproteínas de Membrana/metabolismo , Organelas/fisiologia , Subunidades beta do Complexo de Proteínas Adaptadoras , Animais , Células Cultivadas , Fibroblastos/citologia , Fibroblastos/fisiologia , Biblioteca Genômica , Genótipo , Cor de Cabelo/genética , Proteínas Heterotriméricas de Ligação ao GTP/deficiência , Proteínas Heterotriméricas de Ligação ao GTP/genética , Homozigoto , Glicoproteínas de Membrana Associadas ao Lisossomo , Lisossomos/fisiologia , Lisossomos/ultraestrutura , Melanócitos/citologia , Melanócitos/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monofenol Mono-Oxigenase/metabolismo , Organelas/ultraestrutura , Fenótipo , Subunidades Proteicas , Transporte Proteico
11.
J Vasc Interv Radiol ; 11(9): 1143-52, 2000 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-11041470

RESUMO

PURPOSE: To compare the frequency and extent of pulmonary embolism (PE) occurring during pulse-spray pharmacomechanical thrombolysis (PSPMT) of clotted hemodialysis grafts with use of either urokinase (UK) or heparinized saline (HS). Postintervention primary patency and complication rates were compared for each method of thrombolysis. METHODS AND MATERIALS: Twenty-seven patients were enrolled in this prospective, randomized, double-blind study evaluating PE with two PSPMT agents. The doses of heparin were similar between groups. The only variable was that one group of patients received UK and the other received HS. In two cases, the venous anastomosis could not be crossed. Eleven patients were treated with UK and 14 with HS. Nuclear medicine perfusion lung scans were performed before treatment and after graft declotting procedures. Lung perfusion was quantified to 10% of a pulmonary segment (0 = normal perfusion, 1 = segmental perfusion defect), with nine segments counted for each lung. RESULTS: Baseline nuclear medicine perfusion lung scan results were abnormal (> or = 20% segmental perfusion defect) in 19 patients (70.4%). New PE (one or more pulmonary segments) occurred in two patients treated with UK (18.2%) and nine patients treated with HS (64.3%; P = .04). All cases of PE were asymptomatic. Quantitative global pulmonary perfusion analyses revealed that treatment with UK improved flow to 0.2 +/- 2.0 pulmonary segments, whereas treatment with HS decreased perfusion to 1.0 +/- 1.7 segments (P = .16, NS). Although postintervention primary patency rates were similar according to life-table analysis (P = .76, NS), complication rates were higher with use of HS (n = 4, 28.6%) than with use of UK (n = 2, 18.2%) (P = .6, NS). CONCLUSIONS: All PE were asymptomatic during PSPMT, but treatment with UK reduced the rate of PE and tended to result in smaller defects in lung scan results. Most patients undergoing hemodialysis have abnormal baseline perfusion scan results, but PSPMT with UK improved many of them. The postintervention primary patency rates were similar between groups, but complications were more frequent after treatment with HS.


Assuntos
Prótese Vascular/efeitos adversos , Fibrinolíticos/efeitos adversos , Oclusão de Enxerto Vascular/tratamento farmacológico , Heparina/efeitos adversos , Embolia Pulmonar/etiologia , Diálise Renal/instrumentação , Terapia Trombolítica/efeitos adversos , Ativador de Plasminogênio Tipo Uroquinase/efeitos adversos , Adulto , Idoso , Idoso de 80 Anos ou mais , Derivação Arteriovenosa Cirúrgica/efeitos adversos , Método Duplo-Cego , Feminino , Humanos , Tábuas de Vida , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Embolia Pulmonar/induzido quimicamente , Embolia Pulmonar/prevenção & controle , Radiografia Intervencionista , Resultado do Tratamento , Grau de Desobstrução Vascular
12.
J Am Board Fam Pract ; 13(5): 338-48, 2000.
Artigo em Inglês | MEDLINE | ID: mdl-11001004

RESUMO

BACKGROUND: Although primary care physicians are increasingly interested in adopting electronic medical record (EMR) systems, few use such systems in practice. This study explores the organizational impact of an EMR system on community-based practices that have overcome the initial barriers and are experienced EMR users. METHODS: Five primary care practices that are members of a national research network participated in this study. Using qualitative methods, including semistructured interviews and observations, we assessed the impact of an EMR system on the work lives of various user groups. RESULTS: Physicians and staff indicated that the EMR system has changed not only how they manage patient records but also how they communicate with each other, provide patient care services, and perform job responsibilities. The EMR is also perceived by its users to have an impact on practice costs. Although in most practices physicians and staff were unaware of actual expenses and cost savings associated with the EMR, those in practices that have eliminated duplicate paper-based systems believe they have realized cost savings. CONCLUSIONS: Several important themes emerged. The organizational context in which the system is implemented is important. Effective leadership, the presence of a system champion, availability of technical training and support, and adequate resources are essential elements to the success of the EMR.


Assuntos
Serviços de Saúde Comunitária/organização & administração , Sistemas Computadorizados de Registros Médicos , Atenção Primária à Saúde , Serviços de Saúde Comunitária/economia , Custos de Cuidados de Saúde , Humanos , Atenção Primária à Saúde/economia , Inquéritos e Questionários
13.
Appl Environ Microbiol ; 66(7): 2695-702, 2000 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-10877757

RESUMO

The sorption of organic contaminants by natural organic matter (NOM) often limits substrate bioavailability and is an important factor affecting microbial degradation rates in soils and sediments. We hypothesized that reduced substrate bioavailability might influence which microbial assemblages are responsible for contaminant degradation under enrichment culture conditions. Our primary goal was to characterize enrichments in which different model organic solid phases were used to establish a range of phenanthrene bioavailabilities for soil microorganisms. Phenanthrene sorption coefficients (expressed as log K(D) values) ranged from 3.0 liters kg(-1) for Amberlite carboxylic acid cation-exchange resin (AMB) to 3.5 liters kg(-1) for Biobeads polyacrylic resin (SM7) and 4.2 liters kg(-1) for Biobeads divinyl benzene resin (SM2). Enrichment cultures were established for control (no sorptive phase), sand, AMB, SM7, and SM2 treatments by using two contaminated soils (from Dover, Ohio, and Libby, Mont.) as the initial inocula. The effects of sorption by model phases on the degradation of phenanthrene were evaluated for numerous transfers in order to obtain stable microbial assemblages representative of sorptive and nonsorptive enrichment cultures and to eliminate the effects of the NOM present in the initial inoculum. Phenanthrene degradation rates were similar for each soil inoculum and ranged from 4 to 5 micromol day(-1) for control and sand treatments to approximately 0.4 micromol day(-1) in the presence of the SM7 sorptive phase. The rates of phenanthrene degradation in the highly sorptive SM2 enrichment culture were insignificant; consequently, stable microbial populations could not be obtained. Bacterial isolates obtained from serial dilutions of enrichment culture samples exhibited significant differences in rates of phenanthrene degradation performed in the presence of SM7, suggesting that enrichments performed in the presence of a sorptive phase selected for different microbial assemblages than control treatments containing solid phase phenanthrene.


Assuntos
Bactérias/isolamento & purificação , Bactérias/metabolismo , Fenantrenos/metabolismo , Microbiologia do Solo , Adsorção , Bactérias/crescimento & desenvolvimento , Biodegradação Ambiental , Disponibilidade Biológica , Meios de Cultura , Poluentes do Solo/metabolismo
14.
Appl Environ Microbiol ; 66(7): 2703-10, 2000 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-10877758

RESUMO

Reduced bioavailability of nonpolar contaminants due to sorption to natural organic matter is an important factor controlling biodegradation of pollutants in the environment. We established enrichment cultures in which solid organic phases were used to reduce phenanthrene bioavailability to different degrees (R. J. Grosser, M. Friedrich, D. M. Ward, and W. P. Inskeep, Appl. Environ. Microbiol. 66:2695-2702, 2000). Bacteria enriched and isolated from contaminated soils under these conditions were analyzed by denaturing gradient gel electrophoresis (DGGE) and sequencing of PCR-amplified 16S ribosomal DNA segments. Compared to DGGE patterns obtained with enrichment cultures containing sand or no sorptive solid phase, different DGGE patterns were obtained with enrichment cultures containing phenanthrene sorbed to beads of Amberlite IRC-50 (AMB), a weak cation-exchange resin, and especially Biobead SM7 (SM7), a polyacrylic resin that sorbed phenanthrene more strongly. SM7 enrichments selected for mycobacterial phenanthrene mineralizers, whereas AMB enrichments selected for a Burkholderia sp. that degrades phenanthrene. Identical mycobacterial and Burkholderia 16S rRNA sequence segments were found in SM7 and AMB enrichment cultures inoculated with contaminated soil from two geographically distant sites. Other closely related Burkholderia sp. populations, some of which utilized phenanthrene, were detected in sand and control enrichment cultures. Our results are consistent with the hypothesis that different phenanthrene-utilizing bacteria inhabiting the same soils may be adapted to different phenanthrene bioavailabilities.


Assuntos
Bactérias/genética , Bactérias/metabolismo , Fenantrenos/metabolismo , Microbiologia do Solo , Poluentes do Solo/metabolismo , Adsorção , Bactérias/classificação , Bactérias/isolamento & purificação , Biodegradação Ambiental , Disponibilidade Biológica , Meios de Cultura , DNA Ribossômico/análise , DNA Ribossômico/genética , Eletroforese em Gel de Poliacrilamida/métodos , Genes de RNAr , Dados de Sequência Molecular , Filogenia , Reação em Cadeia da Polimerase , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
15.
Appl Environ Microbiol ; 66(7): 2959-64, 2000 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-10877792

RESUMO

We analyzed the impact of surfactant addition on hydrocarbon mineralization kinetics and the associated population shifts of hydrocarbon-degrading microorganisms in soil. A mixture of radiolabeled hexadecane and phenanthrene was added to batch soil vessels. Witconol SN70 (a nonionic, alcohol ethoxylate) was added in concentrations that bracketed the critical micelle concentration (CMC) in soil (CMC') (determined to be 13 mg g(-1)). Addition of the surfactant at a concentration below the CMC' (2 mg g(-1)) did not affect the mineralization rates of either hydrocarbon. However, when surfactant was added at a concentration approaching the CMC' (10 mg g(-1)), hexadecane mineralization was delayed and phenanthrene mineralization was completely inhibited. Addition of surfactant at concentrations above the CMC' (40 mg g(-1)) completely inhibited mineralization of both phenanthrene and hexadecane. Denaturing gradient gel electrophoresis of 16S rRNA gene segments showed that hydrocarbon amendment stimulated Rhodococcus and Nocardia populations that were displaced by Pseudomonas and Alcaligenes populations at elevated surfactant levels. Parallel cultivation studies revealed that the Rhodococcus population can utilize hexadecane and that the Pseudomonas and Alcaligenes populations can utilize both Witconol SN70 and hexadecane for growth. The results suggest that surfactant applications necessary to achieve the CMC alter the microbial populations responsible for hydrocarbon mineralization.


Assuntos
Bactérias/crescimento & desenvolvimento , Hidrocarbonetos/metabolismo , Microbiologia do Solo , Poluentes do Solo/metabolismo , Tensoativos/metabolismo , Alcanos/química , Alcanos/metabolismo , Bactérias/genética , Bactérias/isolamento & purificação , Bactérias/metabolismo , Biodegradação Ambiental , DNA Bacteriano/genética , Ecossistema , Eletroforese em Gel de Poliacrilamida/métodos , Genes de RNAr , Hidrocarbonetos/química , Fenantrenos/química , Fenantrenos/metabolismo , Reação em Cadeia da Polimerase , RNA Ribossômico 16S/genética
16.
Mol Biol Cell ; 11(7): 2327-33, 2000 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-10888671

RESUMO

Endocytosis in alveolar macrophages can be reversibly inhibited, permitting the isolation of endocytic vesicles at defined stages of maturation. Using an in vitro fusion assay, we determined that each isolated endosome population was capable of homotypic fusion. All vesicle populations were also capable of heterotypic fusion in a temporally specific manner; early endosomes, isolated 4 min after internalization, could fuse with endosomes isolated 8 min after internalization but not with 12-min endosomes or lysosomes. Lysosomes fuse with 12-min endosomes but not with earlier endosomes. Using homogenous populations of endosomes, we have identified Syntaxin 7 as a soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) required for late endosome-lysosome and homotypic lysosome fusion in vitro. A bacterially expressed human Syntaxin 7 lacking the transmembrane domain inhibited homotypic late endosome and lysosome fusion as well as heterotypic late endosome-lysosome fusion. Affinity-purified antibodies directed against Syntaxin 7 also inhibited lysosome fusion in vitro but had no affect on homotypic early endosome fusion. Previous work suggested that human VAMP-7 (vesicle-associated membrane protein-7) was a SNARE required for late endosome-lysosome fusion. A bacterially expressed human VAMP-7 lacking the transmembrane domain inhibited both late endosome-lysosome fusion and homotypic lysosome fusion in vitro. These studies indicate that: 1) fusion along the endocytic pathway is a highly regulated process, and 2) two SNARE molecules, Syntaxin 7 and human VAMP-7, are involved in fusion of vesicles in the late endocytic pathway in alveolar macrophages.


Assuntos
Endossomos/fisiologia , Lisossomos/fisiologia , Macrófagos Alveolares/fisiologia , Proteínas de Membrana/metabolismo , Proteínas de Transporte Vesicular , Animais , Células Cultivadas , Endocitose/fisiologia , Humanos , Macrófagos Alveolares/citologia , Macrófagos Alveolares/metabolismo , Fusão de Membrana/fisiologia , Proteínas de Membrana/genética , Proteínas Qa-SNARE , Proteínas R-SNARE , Coelhos , Ratos , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas SNARE , Solubilidade , Fatores de Tempo
17.
Appl Environ Microbiol ; 66(3): 1038-49, 2000 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-10698769

RESUMO

A variety of contemporary techniques were used to investigate the vertical distribution of thermophilic unicellular cyanobacteria, Synechococcus spp., and their activity within the upper 1-mm-thick photic zone of the mat community found in an alkaline siliceous hot spring in Yellowstone National Park in Wyoming. Detailed measurements were made over a diel cycle at a 61 degrees C site. Net oxygenic photosynthesis measured with oxygen microelectrodes was highest within the uppermost 100- to 200-microm-thick layer until midmorning, but as the day progressed, the peak of net activity shifted to deeper layers, stabilizing at a depth of 300 microm from midday throughout the afternoon. Examination of vertical thin sections by bright-field and autofluorescence microscopy revealed the existence of different populations of Synechococcus which form discrete bands at different vertical positions. Denaturing gradient gel electrophoresis analysis of PCR-amplified 16S rRNA gene segments from horizontal cryosections obtained at 100-microm-thick vertical intervals also suggested vertical stratification of cyanobacterial, green sulfur bacterium-like, and green nonsulfur bacterium-like populations. There was no evidence of diel migration. However, image analysis of vertical thin sections revealed the presence of a narrow band of rod-shaped Synechococcus cells in which the cells assumed an upright position. These upright cells, located 400 to 800 microm below the surface, were observed only in mat samples obtained around noon. In mat samples obtained at other time points, the cells were randomly oriented throughout the mat. These combined observations reveal the existence of a highly ordered structure within the very thin photic zone of this hot spring microbial mat, consisting of morphologically similar Synechococcus populations that are likely to be differentially adapted, some co-occurring with green sulfur bacterium-like populations, and all overlying green nonsulfur bacterium-like populations.


Assuntos
Cianobactérias/citologia , Ecossistema , Temperatura Alta , Microbiologia da Água , Ritmo Circadiano , Cianobactérias/genética , Cianobactérias/efeitos da radiação , Água Doce , Processamento de Imagem Assistida por Computador , Luz , Oxigênio/análise , Fotoperíodo
19.
Environ Microbiol ; 2(4): 428-35, 2000 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-11234931

RESUMO

Inferences about the evidence of life recorded in organic compounds within the Earth's ancient rocks have depended on 13C contents low enough to be characteristic of biological debris produced by the well-known CO2 fixation pathway, the Calvin cycle. 'Atypically' high values have been attributed to isotopic alteration of sedimentary organic carbon by thermal metamorphism. We examined the possibility that organic carbon characterized by a relatively high 13C content could have arisen biologically from recently discovered autotrophic pathways. We focused on the green non-sulphur bacterium Chloroflexus aurantiacus that uses the 3-hydroxypropionate pathway for inorganic carbon fixation and is geologically significant as it forms modern mat communities analogous to stromatolites. Organic matter in mats constructed by Chloroflexus spp. alone had relatively high 13C contents (-14.9%) and lipids diagnostic of Chloroflexus that were also isotopically heavy (-8.9% to -18.5%). Organic matter in mats constructed by Chloroflexus in conjunction with cyanobacteria had a more typical Calvin cycle signature (-23.5%). However, lipids diagnostic of Chloroflexus were isotopically enriched (-15.1% to -24.1%) relative to lipids typical of cyanobacteria (-33.9% to -36.3%). This suggests that, in mats formed by both cyanobacteria and Chloroflexus, autotrophy must have a greater effect on Chloroflexus carbon metabolism than the photoheterotrophic consumption of cyanobacterial photosynthate. Chloroflexus cell components were also selectively preserved. Hence, Chloroflexus autotrophy and selective preservation of its products constitute one purely biological mechanism by which isotopically heavy organic carbon could have been introduced into important Precambrian geological features.


Assuntos
Isótopos de Carbono/metabolismo , Chlorobi/metabolismo , Cianobactérias/metabolismo , Água Doce/microbiologia , Compostos Orgânicos/metabolismo , Biofilmes/crescimento & desenvolvimento , Chlorobi/crescimento & desenvolvimento , Cromatografia Gasosa/métodos , Cianobactérias/crescimento & desenvolvimento , Lipídeos/análise , Pigmentos Biológicos/análise
20.
Traffic ; 1(11): 816-22, 2000 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-11208072

RESUMO

Chediak-Higashi syndrome (CHS) is a rare autosomal recessive disorder of human, mouse (beige) and other mammalian species. The same genetic defect was found to result in the disease in all species identified, permitting a positional cloning approach using the mouse model beige to identify the responsible gene. The CHS gene was cloned and mutations identified in affected species. This review discusses the clinical features of CHS contrasting features seen in similar syndromes. The possible functions of the protein encoded by the CHS/beige gene are discussed, along with the alterations in cellular physiology seen in mutant cells.


Assuntos
Síndrome de Chediak-Higashi/genética , Animais , Síndrome de Chediak-Higashi/diagnóstico , Síndrome de Chediak-Higashi/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Metabolismo dos Lipídeos , Lisossomos/patologia , Camundongos , Camundongos Mutantes , Mutação , Proteínas/genética , Transdução de Sinais , Linfócitos T Citotóxicos/patologia , Proteínas de Transporte Vesicular
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