Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 36
Filtrar
1.
Nat Commun ; 9(1): 1897, 2018 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-29765044

RESUMO

Complex-oxide materials exhibit physical properties that involve the interplay of charge and spin degrees of freedom. However, an ambipolar oxide that is able to exhibit both electron-doped and hole-doped ferromagnetism in the same material has proved elusive. Here we report ambipolar ferromagnetism in LaMnO3, with electron-hole asymmetry of the ferromagnetic order. Starting from an undoped atomically thin LaMnO3 film, we electrostatically dope the material with electrons or holes according to the polarity of a voltage applied across an ionic liquid gate. Magnetotransport characterization reveals that an increase of either electron-doping or hole-doping induced ferromagnetic order in this antiferromagnetic compound, and leads to an insulator-to-metal transition with colossal magnetoresistance showing electron-hole asymmetry. These findings are supported by density functional theory calculations, showing that strengthening of the inter-plane ferromagnetic exchange interaction is the origin of the ambipolar ferromagnetism. The result raises the prospect of exploiting ambipolar magnetic functionality in strongly correlated electron systems.

2.
Zhonghua Yan Ke Za Zhi ; 53(2): 104-108, 2017 Feb 11.
Artigo em Chinês | MEDLINE | ID: mdl-28260360

RESUMO

Objective: To evaluate the usefulness of albumin correction in determination of cytomegalovirus IgG in the aqueous humor of Posner-Schlossman syndrome (PSS) patients. Methods: Cases series studies. Forty-two patients (26 men and 16 women) who were diagnosed as PSS were enrolled from Oct. 2009 to Oct. 2015 at the Eye and ENT Hospital. During the same period, 20 patients with primary open-angle glaucoma (POAG) and 30 patients with bacterial endophthalmitis or retinal necrosis were enrolled as negative control group and inflammatory disease control group, respectively. Aqueous humor and serum samples were assayed to detect CMV IgG by enzyme-linked immunosorbent assay (ELISA), and albumin by scattering immunonephelometry. CMV DNA in aqueous humor was assayed by polymerase chain reaction (PCR). The ratio which was calculated as the (aqueous humor CMV IgG/serum CMV IgG)/(aqueous humor concentration of albumin/serum albumin concentration) over 0.6 was considered as intraocular antibody formation. Performance of differentiating control eyes from eyes with CMV-positive PSS was evaluated by the receiver operating characteristic curve. The ANOVA test, Mann-Whitney test and Chi-square test were performed to compare the differences among groups. Results: The detectable rate of CMV IgG antibody in the aqueous humor was 76.2%, 100.0% and 10.0% in PSS, inflammatory disease control and POAG groups, respectively. The levels of CMV IgG antibody in the PSS groups were significantly higher than that of POAG groups (Z=4.23, P<0.001).The positive rate corrected by the albumin was 71.4%, 3.3% and 0.0%.The corrected positive rate in PSS groups was significantly higher than that of the inflammatory disease control and POAG groups (χ(2)=30.38, P<0.01; χ(2)= 24.89, P<0.01), with a sensitivity of 75.0% and a specificity of 98.0%. The area under the curve for calibrated ratio was 0.942 (95%CI: 0.859 to 0.984) which was higher than that of CMV IgG (Z=6.19, P<0.001).The corrected positive rate of CMV IgG antibody (71.4%) was higher than that of CMV DNA (47.6%, χ(2)=4.003, P=0.045). Conclusions: CMV IgG antibody ratio which was corrected by aqueous humor and serum albumin could effectively improve aqueous antibody specificity in PSS patients. Furthermore, CMV IgG antibody ratio combined with PCR could improve the sensitivity of CMV detection. All of which help clarify the CMV infection in PSS in CMV DNA negative eyes. (Chin J Ophthalmol, 2017, 53: 104-108).


Assuntos
Albuminas/análise , Humor Aquoso/imunologia , Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Imunoglobulina G/análise , Iridociclite/imunologia , Hipertensão Ocular/imunologia , Estudos de Casos e Controles , Citomegalovirus/genética , DNA Viral/análise , Endoftalmite/imunologia , Endoftalmite/microbiologia , Ensaio de Imunoadsorção Enzimática , Feminino , Glaucoma de Ângulo Aberto/imunologia , Humanos , Imunoglobulina G/sangue , Masculino , Necrose , Hipertensão Ocular/virologia , Reação em Cadeia da Polimerase , Retina/patologia , Albumina Sérica/análise , Síndrome
3.
Nanoscale ; 9(8): 2706-2710, 2017 Feb 23.
Artigo em Inglês | MEDLINE | ID: mdl-28191573

RESUMO

Photodynamic therapy (PDT) is a non-invasive therapy with many advantages over other therapeutic methods, but it is restricted to treat superficial cancers due to the shallow tissue penetration of visible light. The biological window in the near infrared region (NIR) offers hope to extend the penetration depth, but there is no natural NIR excited photosensitizer. Here, we report a novel photosensitizer: NaYbF4 nanoparticles (NPs). By using a 1,3-diphenylisobenzofuran (DPBF) sensor, we show that the Yb3+ ions can absorb the NIR light and transfer energy directly to oxygen to generate reactive oxygen species (ROS). The efficiency of transferring energy to oxygen by NaYbF4 NPs is comparable to that of traditional photosensitizers. We have carried out PDT both in vitro and in vivo based on NaYbF4 NPs; the results demonstrate that NaYbF4 NPs are indeed an effective NIR photosensitizer, which can help extend the application of PDT to solid tumors owing to the much deeper penetration depth of NIR light.


Assuntos
Raios Infravermelhos , Nanopartículas , Neoplasias/tratamento farmacológico , Fotoquimioterapia , Fármacos Fotossensibilizantes/química , Animais , Linhagem Celular Tumoral , Humanos , Camundongos Nus
4.
Zhonghua Yan Ke Za Zhi ; 52(2): 123-8, 2016 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-26906708

RESUMO

OBJECTIVE: To determine the success rates and compare the results of balloon catheter dilation and nasolacrimal intubation as treatment for congenital nasolacrimal duct obstruction after failed probing, stratified by category of age and type of obstruction. METHODS: It was a prospective, randomized, clinical trial that enrolled 189 children (245 eyes) aged between 6 months to 48 months who had a history of failed nasolacrimal duct probing. All eyes underwent either balloon catheter nasolacrimal duct dilation or nasolacrimal duct intubation randomly. The eyes were divided into 2 age categories: category 1 (6-24 months) and category 2 (>24 months) and into 2 types of obstructions: simple obstruction and complex obstruction. Treatment success was defined as absence of epiphora, mucous discharge, or increased lacrimal lake at the outcome visit 6 months after surgery. Complications were also compared. RESULTS: In 124 eyes treated with balloon catheter dilatation, 112 were successful (90.3%) comparing with 106 successful eyes (87.6%) in 121 eyes treated with nasolacrimal duct intubation. The risk ratio for success between intubation and balloon dilation was 0.971, and the 95% confidence interval was 0.95-1.22. Within each age category, the success rate varied but did not show significant difference: In those under 24 months, success rate was 89.7% in 97 eyes treated with intubation, and 91.9% in 99 eyes treated with balloon dilation (RR, 0.976; 95% CI, 0.590-0.956). In those above 24 months, success rate was 79.1% in 24 eyes treated with intubation, and 84.0% in 25 eyes treated with balloon dilation (RR, 0.942; 95%CI, 0.813-1.387). In the group of simple obstruction, success rate was 96.5% in 87 eyes treated with intubation, and 93.1% in 88 eyes treated with balloon dilation (RR, 1.036; 95% CI, 0.967-1.105). In the group of complex obstruction, Success rate was 64.7% in 34 eyes treated with intubation, and 86.1% in 36 eyes treated with balloon dilation. The success rate of balloon dilatation showed slightly higher than that of intubation (RR, 0.751; 95% CI, 0.590-0.956). There were 59 eyes showed complications in intubation group, while only 2 eyes in balloon dilation group. CONCLUSIONS: Both balloon catheter dilation and nasolacrimal duct intubation could alleviate the clinical signs of persistent nasolacrimal duct obstruction with a similar percentage of patients. In the complex obstruction group, balloon catheter dilation showed better efficacy than nasolacrimal duct intubation.


Assuntos
Dilatação/métodos , Intubação/métodos , Obstrução dos Ductos Lacrimais/congênito , Obstrução dos Ductos Lacrimais/terapia , Silício , Pré-Escolar , Humanos , Lactente , Obstrução dos Ductos Lacrimais/classificação , Ducto Nasolacrimal , Estudos Prospectivos , Resultado do Tratamento
5.
Genet Mol Res ; 14(3): 9604-13, 2015 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-26345893

RESUMO

We investigated the killing effect of low-intensity ultrasound combined with 5-aminolevulinic acid (5-ALA) on the rat osteosarcoma cell line UMR-106. Logarithmic-phase UMR-106 cells were divided into a control group, ultrasound group and 5-ALA group. The cell apoptotic rate, production of reactive oxygen species, and the change in mitochondrial membrane potential were analyzed by flow cytometry; ultrastructural changes were observed by transmission electron microscopy. Using low-intensity ultrasound at 1.0 MHz and 2.0 W/cm(2) plus 5-ALA at a concentration of 2 mM, the apoptotic rate of the sonodynamic therapy group was 27.2 ± 3.4% which was significantly higher than that of the control group, ultrasound group, and 5-ALA group (P < 0.05). The production of reactive oxygen species was 32.6 ± 2.2% and the decrease in mitochondrial membrane potential was 39.5 ± 2.5%. The 33342 staining showed nuclear condensation and fragmentation in the ultrasound group and 5-ALA group. Structural changes in the cell membrane, mitochondria, Golgi apparatus, and other organelles observed by transmission electron microscopy included formation of apoptotic bodies. The killing effect of low-intensity ultrasound combined with 5-ALA on UMR-106 cells was significant. Cell apoptosis played a vital role in the killing effect, and the mitochondria pathway contributed to the apoptosis of UMR-106 cells.


Assuntos
Ácido Aminolevulínico/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Ondas Ultrassônicas/efeitos adversos , Linhagem Celular Tumoral , Citometria de Fluxo , Humanos , Osteossarcoma/metabolismo , Osteossarcoma/terapia , Espécies Reativas de Oxigênio/metabolismo , Terapia por Ultrassom
6.
Appl Environ Microbiol ; 67(8): 3577-85, 2001 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-11472934

RESUMO

Mycobacterium sp. strain PYR-1 degrades high-molecular-weight polycyclic hydrocarbons (PAHs) primarily through the introduction of both atoms of molecular oxygen by a dioxygenase. To clone the dioxygenase genes involved in PAH degradation, two-dimensional (2D) gel electrophoresis of PAH-induced proteins from cultures of Mycobacterium sp. strain PYR-1 was used to detect proteins that increased after phenanthrene, dibenzothiophene, and pyrene exposure. Comparison of proteins from induced and uninduced cultures on 2D gels indicated that at least six major proteins were expressed (105, 81, 52, 50, 43, and 13 kDa). The N-terminal sequence of the 50-kDa protein was similar to those of other dioxygenases. A digoxigenin-labeled oligonucleotide probe designed from this protein sequence was used to screen dioxygenase-positive clones from a genomic library of Mycobacterium sp. strain PYR-1. Three clones, each containing a 5,288-bp DNA insert with three genes of the dioxygenase system, were obtained. The genes in the DNA insert, from the 5' to the 3' direction, were a dehydrogenase, the dioxygenase small (beta)-subunit, and the dioxygenase large (alpha)-subunit genes, arranged in a sequence different from those of genes encoding other bacterial dioxygenase systems. Phylogenetic analysis showed that the large alpha subunit did not cluster with most of the known alpha-subunit sequences but rather with three newly described alpha subunits of dioxygenases from Rhodococcus spp. and Nocardioides spp. The genes from Mycobacterium sp. strain PYR-1 were subcloned and overexpressed in Escherichia coli with the pBAD/ThioFusion system. The functionality of the genes for PAH degradation was confirmed in a phagemid clone containing all three genes, as well as in plasmid subclones containing the two genes encoding the dioxygenase subunits.


Assuntos
Clonagem Molecular , Mycobacterium/enzimologia , Oxigenases/genética , Oxigenases/metabolismo , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Dados de Sequência Molecular , Mycobacterium/genética , Filogenia , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Análise de Sequência de DNA
7.
Appl Environ Microbiol ; 66(10): 4300-4, 2000 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-11010873

RESUMO

A 81-kDa protein from Mycobacterium sp. strain PYR-1 was expressed in response to exposure of the strain to the polycyclic aromatic hydrocarbon pyrene and recovered by two-dimensional gel electrophoresis. The N-terminal sequence of the protein indicated that it was similar to catalase-peroxidase. An oligonucleotide probe designed from this sequence was used to screen a genomic library of Mycobacterium sp. strain PYR-1, and a positive clone, containing a part of the gene encoding the 81-kDa protein, was isolated. A gene-walking technique was used to sequence the entire gene, which was identified as katG for catalase-peroxidase. The deduced KatG protein sequence showed significant homology to KatGII of Mycobacterium fortuitum and clustered with catalase-peroxidase proteins from other Mycobacterium species in a phylogenetic tree. The katG gene was expressed in Escherichia coli to produce a protein with catalase-peroxidase activity. Since the induction of this catalase-peroxidase occurred in pyrene-induced cultures and the exposure of these cultures to hydrogen peroxide reduced pyrene metabolism, our data suggest that this enzyme plays a role in polycyclic aromatic hydrocarbon metabolism by strain PYR-1.


Assuntos
Proteínas de Bactérias , Mycobacterium/enzimologia , Mycobacterium/genética , Peroxidases/genética , Peroxidases/metabolismo , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Sequência de Bases , Clonagem Molecular , Escherichia coli/genética , Vetores Genéticos , Peróxido de Hidrogênio/farmacologia , Cinética , Dados de Sequência Molecular , Mycobacterium/efeitos dos fármacos , Filogenia , Plasmídeos , Pirenos/metabolismo , Proteínas Recombinantes/metabolismo
8.
FEMS Microbiol Lett ; 188(1): 55-61, 2000 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-10867234

RESUMO

A polyclonal antibody against microsomes of a fungus, Cunninghamella elegans, was used to screen a C. elegans cDNA library. A cDNA clone, containing an open reading frame (ORF) encoding a protein of 389 amino acids (aa), was obtained. GenBank comparison (BLAST) showed that the protein was closely related to P450 because a heme-binding region, which is highly conserved in all P450 sequences, was found in the ORF protein. Using an oligo probe designed from this C. elegans heme-binding region to rescreen the cDNA library, we obtained three new clones. Sequence comparison showed that the three clones, with different length cDNA inserts, were from the same mRNA of the C. elegans P450 gene. One clone had the full C. elegans P450 gene, encoding 473 aa with a molecular mass of 54958.60, whereas the 389 was a part of the 473 aa without the N-terminal. The entire C. elegans P450 gene was successfully subcloned and overexpressed in a plasmid-Escherichia coli system (pQE30). Immunostaining with three antibodies (CYP1A1, CYP2E1, and CYP3A1) against mammalian P450 enzymes and benzidine staining for hemoproteins showed positive results for the recombinant protein expressed in E. coli. A phylogenetic tree was constructed by comparison of other fungal P450s to the C. elegans sequence. The C. elegans P450 clustered close to the cyp51 family and was named cyp509A1 by the International Committee on the Nomenclature for Cytochrome P450 Enzymes.


Assuntos
Cunninghamella/enzimologia , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Escherichia coli/genética , Sequência de Aminoácidos , Sequência de Bases , Clonagem Molecular , Cunninghamella/genética , Sistema Enzimático do Citocromo P-450/química , Eletroforese em Gel de Poliacrilamida , Escherichia coli/enzimologia , Dados de Sequência Molecular , Filogenia , Análise de Sequência de DNA
9.
Mol Cell Probes ; 14(1): 1-5, 2000 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-10722787

RESUMO

Rapid identification of bovine materials in animal feedstuffs is essential for effective control of a potential source of bovine spongiform encephalopathy. We have developed a rapid method for the detection of the presence of bovine materials in animal feeds. Animal feed samples were prepared by a Chelex-100 treatment method, then subjected to polymerase chain reaction (PCR) detection. The assay can be completed in 2 h including 30 min for sample preparation, 35-65 min for PCR cycling and 30 min for gel electrophoresis. This method is not only rapid, simple and consistent, but also avoids a hazardous waste disposal issue associated with a previously described guanidine thiocyanate (GuSCN) extraction-PCR method.


Assuntos
Ração Animal/normas , Reação em Cadeia da Polimerase/métodos , Ração Animal/análise , Animais , Produtos Biológicos , Bovinos/genética , Primers do DNA , Encefalopatia Espongiforme Bovina/prevenção & controle , Encefalopatia Espongiforme Bovina/transmissão , Carne/análise , Minerais/análise , Reação em Cadeia da Polimerase/instrumentação , Sensibilidade e Especificidade , Fatores de Tempo
10.
Mol Cell Probes ; 13(1): 29-33, 1999 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-10024430

RESUMO

Mouse hepatitis virus (MHV) infection in laboratory mouse populations is a serious problem, because the MHV infections are known to interfere with research results. Confirmation of indirect serological detection methods by viral isolation is difficult. Reverse transcription plus polymerase chain reaction (RT-PCR) was used to test 94 mouse tissue samples from suspected naturally MHV infected mice. Positive results were only obtained from two colon samples and one mixed sample with colon and liver. The low positive rate is probably due to the virus being rapidly cleared by the MHV antibodies produced in the mouse. However, RT-PCR detection of MHV in nude mice placed in the same cages with other non-nude mice or placed in cages with used dirty bedding, showed a very high positive rate: 10 out of 12 colon samples were positive (83%), and 5 out of 10 faecal samples were positive (50%). A single-tube, single step RT-PCR method and two procedures for isolation of the viral RNA for the RT-PCR assay were also included in this article.


Assuntos
Infecções por Coronavirus/veterinária , Hepatite Viral Animal/diagnóstico , Vírus da Hepatite Murina/isolamento & purificação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Animais , Colo/virologia , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/virologia , Fezes/virologia , Hepatite Viral Animal/virologia , Abrigo para Animais , Fígado/virologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Vírus da Hepatite Murina/genética , RNA Viral/isolamento & purificação , Vigilância de Evento Sentinela
11.
FEMS Microbiol Lett ; 169(2): 397-402, 1998 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-9868787

RESUMO

The fungus, Cunninghamella elegans has been widely used in bioremediation and microbial models of mammalian studies in many laboratories. Using the polymerase chain reaction to randomly amplify the insert directly from the single non-blue plaques of a C. elegans cDNA library, then partly sequencing and comparing with GenBank sequences, we have identified a clone which contains C. elegans 6-phosphogluconate dehydrogenase gene. The polymerase chain reaction product was cloned into a plasmid, pGEM-T Easy vector for full insert DNA sequencing. The 6-phosphogluconate dehydrogenase gene (1458 bases) and the deduced protein sequence were determined from the insert DNA sequence. The gene was found by open reading frame analysis and confirmed by the alignment of the deduced protein sequence with other published 6-phosphogluconate dehydrogenase sequences. Several highly conserved regions were found for the 6-phosphogluconate dehydrogenase sequences. The 6-phosphogluconate dehydrogenase gene was subcloned and over-expressed in a plasmid-E. coli system (pQE30). The cell lysate of this clone has a very high 6-phosphogluconate dehydrogenase enzyme activity. Most of the recombinant protein in this system was formed as insoluble inclusion bodies, but soluble in high concentration of urea-buffer. Ni-NTA resin was used to purify the recombinant protein which showed 6-phosphogluconate dehydrogenase enzyme activity. The recombinant protein has a predicted molecular size correlating with that revealed by sodium dodecylsulfate-polyacrylamide gel electrophoresis analysis. The C. elegans 6-phosphogluconate dehydrogenase was in a cluster with yeast' 6-phosphogluconate dehydrogenase in the phylogenetic tree. Bacterial 6-phosphogluconate dehydrogenase and higher organisms' 6-phosphogluconate dehydrogenase were found in different clusters.


Assuntos
Cunninghamella/genética , DNA Complementar/genética , Proteínas Fúngicas/genética , Fosfogluconato Desidrogenase/genética , Sequência de Aminoácidos , Clonagem Molecular , Cunninghamella/enzimologia , Eletroforese em Gel de Poliacrilamida , Proteínas Fúngicas/biossíntese , Genes Bacterianos , Vetores Genéticos , Dados de Sequência Molecular , Fases de Leitura Aberta , Fosfogluconato Desidrogenase/biossíntese , Reação em Cadeia da Polimerase , Proteínas Recombinantes/biossíntese , Alinhamento de Sequência/classificação
12.
J Immunol Methods ; 215(1-2): 123-34, 1998 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-9744754

RESUMO

Changes in the patterns of cytokine expression are thought to be of central importance in human infectious and inflammatory diseases. As such, there is a need for precise, reproducible assays for quantification of cytokine mRNA that are amenable to routine use in a clinical setting. In this report, we describe the design and performance of a branched DNA (bDNA) assay for the direct quantification of multiple cytokine mRNA levels in peripheral blood mononuclear cells (PBMCs). Oligonucleotide target probe sets were designed for several human cytokines, including TNFalpha, IL-2, IL-4, IL-6, IL-10, and IFNgamma. The bDNA assay yielded highly reproducible quantification of cytokine mRNAs, exhibited a broad linear dynamic range of over 3-log10, and showed a sensitivity sufficient to measure at least 3000 molecules. The potential clinical utility of the bDNA assay was explored by measuring cytokine mRNA levels in PBMCs from healthy and immunocompromised individuals. Cytokine expression levels in PBMCs from healthy blood donors were found to remain relatively stable over a one-month period of time. Elevated levels of IFNgamma mRNA were detected in PBMCs from HIV-1 seropositive individuals, but no differences in mean levels of TNFalpha or IL-6 mRNA were detected between seropositive and seronegative individuals. By providing a reproducible method for quantification of low abundance transcripts in clinical specimens, the bDNA assay may be useful for studies addressing the role of cytokine expression in disease.


Assuntos
Citocinas/biossíntese , DNA/análise , Leucócitos Mononucleares/metabolismo , RNA Mensageiro/sangue , Citocinas/sangue , Soropositividade para HIV/sangue , HIV-1/imunologia , Humanos , Modelos Lineares , Conformação de Ácido Nucleico , Sondas de Oligonucleotídeos , Reprodutibilidade dos Testes , Sensibilidade e Especificidade
13.
Exp Gerontol ; 33(3): 239-47, 1998 May.
Artigo em Inglês | MEDLINE | ID: mdl-9615922

RESUMO

The effect of a food-restricted diet on the fecal microflora of rats was studied by determining total anaerobic bacteria, bacterial cellular fatty acids, and the predominant intestinal bacteria shown by polymerase chain reaction (PCR) primers specific for the 16S rRNA gene sequences of 12 bacterial species. Twenty-four female Fischer 344 rats, 57 days old were divided into two groups and maintained on an NIH-31 diet. One group was fed ad libitum while the other group received 60% of ad libitum food intake (40% food restriction supplemented with vitamins and minerals equal to the ad libitum animals). After 2, 10, and 20 weeks on this dietary regimen, groups of four animals were sacrificed and the intestinal contents analyzed for changes in the bacterial flora. The anaerobic population for two-week (short-term) food-restricted rats was 3.2 x 10(8) per gram, slightly less than the 9.1 x 10(8) per gram found in the ad libitum-fed rats. The anaerobic populations in 20-week food restricted and ad libitum fed rats were 1.9 x 10(9) and 2.7 x 10(9) per gram, respectively. The total anaerobic population did not change significantly in either group during the 20-week study. No statistically significant differences were observed in the bacterial cellular fatty acid profiles between the two groups as determined by gas-liquid chromatography. PCR analysis of the intestinal contents indicated no significant shifts in the predominant flora due to dietary changes. The results, using three different methods to detect changes in the rat intestinal microflora, suggest that long-term dietary restriction had little effect on the microflora of female Fischer 344 rats.


Assuntos
Privação de Alimentos/fisiologia , Intestinos/microbiologia , Animais , Bactérias Anaeróbias/genética , Bactérias Anaeróbias/crescimento & desenvolvimento , Bactérias Anaeróbias/metabolismo , Contagem de Colônia Microbiana , Ácidos Graxos/metabolismo , Fezes/microbiologia , Feminino , Reação em Cadeia da Polimerase , RNA Ribossômico 16S/análise , Ratos , Ratos Endogâmicos F344
14.
Mol Cell Probes ; 11(4): 259-65, 1997 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-9281411

RESUMO

Polymerase chain reaction (PCR) procedures for Ruminococcus spp. were developed and used for the detection of Ruminococcus spp. in human and animal faeces. The PCR specificity was established for Ruminococcus albus, R. bromii, R. obeum, R. callidus and R. flavefaciens. The PCR sensitivity ranged from 4 to 100 cells of the pure cultures, depending on the species. Ruminococcus albus, R. bromii, R. obeum and R. callidus were detected in human faeces. R. bromii, R. albus and R. obeum were also detected in animal faeces. R. flavefaciens was not detected in most of the samples tested, except for a weak positive in monkey faecal specimens. The PCR appears to be a specific and efficient method for the detection of Ruminococcus spp. in faecal specimens.


Assuntos
Fezes/microbiologia , Cocos Gram-Positivos/genética , Reação em Cadeia da Polimerase/métodos , Animais , Bactérias Anaeróbias/classificação , Bactérias Anaeróbias/genética , Técnicas de Tipagem Bacteriana , Cocos Gram-Positivos/classificação , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Ratos , Ratos Endogâmicos F344 , Sensibilidade e Especificidade , Especificidade da Espécie
15.
Mol Cell Probes ; 11(6): 427-32, 1997 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-9500811

RESUMO

Oligonucleotide probes were used for identification of Shigella and analysis of the relationship between Shigella spp. and Escherichia coli. Probe-based PCRs shown cross-reactions from Shigella to E. coli. Probe-based 16S rRNA sequencing and phylogenetic analysis showed the four species of Shigella: Sh. dysenteriae, Sh. boydii, Sh. sonnei, and Sh. flexneri, formed a cluster with E. coli. Shigella flexneri and Sh. sonnei are even more similar to E. coli than to the other two Shigella species. These results confirmed an earlier recommendation that the four species of Shigella and E. coli should be classified as five sub-groups within a single species.


Assuntos
Filogenia , Reação em Cadeia da Polimerase/métodos , RNA Ribossômico 16S/genética , Shigella/genética , Sondas de DNA , Escherichia coli/genética , Amplificação de Genes , Análise de Sequência de DNA
16.
J Appl Microbiol ; 83(6): 727-36, 1997 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-9449811

RESUMO

A universal protocol for PCR detection of 13 species of foodborne pathogens in foods was developed. The protocol used a universal culture medium and the same PCR conditions with 13 sets of specific primers. The 13 species of foodborne pathogens examined were Escherichia coli, E. coli-ETEC, E. coli-O157:H7, Shigella spp., Salmonella spp., Yersinia enterocolitica, Y. pseudotuberculosis, Vibrio cholerae, V. parahaemolyticus, V. vulnificus, Listeria monocytogenes, Staphylococcus aureus and Bacillus cereus. No interference was observed using the PCR assay when food sample was artificially inoculated with each individual bacterial species. Twelve different seafood samples and two soft cheese samples without artificial inoculation were examined by this protocol. Vibrio vulnificus, Salmonella spp., E. coli, Listeria monocytogenes and Bacillus cereus were detected in some foods. Internal probe hybridization and nested PCR procedures were used to confirm the above findings.


Assuntos
Bactérias/isolamento & purificação , Microbiologia de Alimentos , Reação em Cadeia da Polimerase , Alimentos Marinhos/microbiologia
18.
Appl Environ Microbiol ; 62(5): 1825-30, 1996 May.
Artigo em Inglês | MEDLINE | ID: mdl-8633883

RESUMO

The 2,3-dihydroxybiphenyl 1,2-dioxygenase (2,3-DBPD) of Pseudomonas putida OU83 was constitutively expressed and purified to apparent homogeneity. The apparent molecular mass of the native enzyme was 256 kDa, and the subunit molecular mass was 32 kDa. The data suggested that 2,3-DBPD was an octamer of identical subunits. The nucleotide sequence of a DNA fragment containing the bphC region was determined. The deduced protein sequence for 2,3-DBPD consisted of 292 amino acid residues, with a calculated molecular mass of 31.9 kDa, which was in agreement with data for the purified 2,3-DBPD. Nucleotide and amino acid sequence analyses of the bphC gene and its product, respectively, revealed that there was a high degree of homology between the OU83 bphC gene and the bphC genes of Pseudomonas cepacia LB400 and Pseudomonas pseudoalcaligenes KF707.


Assuntos
Oxirredutases/isolamento & purificação , Pseudomonas putida/enzimologia , Sequência de Aminoácidos , Sequência de Bases , Dados de Sequência Molecular , Oxirredutases/genética , Filogenia , Pseudomonas putida/genética , Alinhamento de Sequência , Análise de Sequência
19.
Appl Environ Microbiol ; 62(4): 1242-7, 1996 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-8919784

RESUMO

PCR procedures based on 16S rRNA gene sequences specific for 12 anaerobic bacteria that predominate in the human intestinal tract were developed and used for quantitative detection of these species in human (adult and baby) feces and animal (rat, mouse, cat, dog, monkey, and rabbit) feces. Fusobacterium prausnitzii, Peptostreptococcus productus, and Clostridium clostridiiforme had high PCR titers (the maximum dilutions for positive PCR results ranged from 10(-3) to 10(-8)) in all of the human and animal fecal samples tested. Bacteroides thetaiotaomicron, Bacteroides vulgatus, and Eubacterium limosum also showed higher PCR titers (10(-2) to 10(-6)) in adult human feces. The other bacteria tested, including Escherichia coli, Bifidobacterium adolescentis, Bifidobacterium longum, Lactobacillus acidophilus, Eubacterium biforme, and Bacteroides distasonis, were either at low PCR titers (less than 10(-2)) or not detected by PCR. The reported PCR procedure including the fecal sample preparation method is simplified and rapid and eliminates the DNA isolation steps.


Assuntos
Bactérias Anaeróbias/genética , Bactérias Anaeróbias/isolamento & purificação , Fezes/microbiologia , Reação em Cadeia da Polimerase/métodos , Adulto , Animais , Sequência de Bases , Gatos , Primers do DNA/genética , DNA Bacteriano/genética , Cães , Estudos de Avaliação como Assunto , Humanos , Lactente , Camundongos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase/estatística & dados numéricos , RNA Bacteriano/genética , RNA Ribossômico 16S/genética , Coelhos , Ratos , Sensibilidade e Especificidade
20.
Int J Syst Bacteriol ; 46(2): 466-9, 1996 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-8934904

RESUMO

A bacterium isolated from a polluted stream, capable of metabolizing biphenyl, naphthalene, phenanthrene, and higher-molecular-weight polycyclic aromatic hydrocarbons (D. Gibson, V. Mahadevan, D. Jerina, H. Yagi, and H. Yeh, Science 189:295-297, 1975), was previously identified as Beijerinckia sp. strain B1. In this investigation, 16S rRNA gene sequencing, biochemical tests, fatty acid methyl ester analysis, polyacrylamide gel electrophoresis of protein, and DNA-DNA hybridization were used to determine the taxonomic relationship of Beijerinckia sp. strain B1. The sequence of the 16S rRNA gene of B1 was identical to that of Sphingomonas yanoikuyae ATCC 51230T. The biochemical tests, fatty acid analysis, and sodium dodecyl sulfate-polacrylamide gel electrophoresis profile of soluble proteins of strain B1 showed results similar to those of S. yanoikuyae. DNA-DNA hybridization indicated that B1 and S. yanoikuyae ATCC 51230T are 75% homologous at the DNA level. We propose that Beijerinckia sp. strain B1 be reclassified as S. yanoikuyae.


Assuntos
Ácidos Graxos/análise , Bactérias Aeróbias Gram-Negativas/classificação , Microbiologia do Solo , Proteínas de Bactérias/análise , Técnicas de Tipagem Bacteriana , DNA Bacteriano/análise , Eletroforese em Gel de Poliacrilamida , Ésteres/análise , Bactérias Aeróbias Gram-Negativas/enzimologia , Bactérias Aeróbias Gram-Negativas/genética , Hidrocarbonetos/metabolismo , Hibridização de Ácido Nucleico , Fenótipo , Filogenia , RNA Ribossômico 16S/genética
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...